Vectra Polaris
User Manual
TM
P/N CLS147553 Rev. G
Publication Date: July 27, 2018
Instrument Safety
The following safety information about the Vectra Polaris is included
in this documentation. Read and review all safety information
before operating the Vectra Polaris.
• Required Training
• Electrical Safety
• Mechanical Safety
• Weight Warning
• Bright Light
Required Training
Ensure that all personnel involved with the operation of the
instrument have:
• Received instruction in general safety practices for laboratories.
• Received instruction in specific safety practices for the
instrument.
Instrument Safety 2
WARNING
Use this product only in the manner described in this manual. If the
equipment is used in a manner not specified by the manufacturer,
the protection provided by the equipment may be impaired.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Electrical Safety
The Vectra Polaris is powered by a 100-120VAC/200-240VAC, 5060Hz (±10%) input power supply.
The wall outlet or the power cable connector on the left side of the
instrument should be accessible after the system's installation, to
enable trained service personnel to safely disconnect power from
the system during servicing.
WARNING
Do not operate the system in an environment with explosive or
flammable gases.
WARNING
Instrument Safety 3
• DO NOT remove instrument covers. There are no user
serviceable parts inside. The covers are intended to be removed
only by qualified PerkinElmer service personnel; they are not
intended to be removed during operation or for maintenance by
users. Contact PerkinElmer technical support if help is required
(see page 7).
• Do not operate the system if there has been a malfunction of the
system door or slide loading components. Contact PerkinElmer
technical support if help is required (see page 7).
• Do not operate the system in places where it may be splashed
with liquid.
For further electrical safety information, refer to the following
sections:
• Power Cord Selection
• Fuses
• Cables and Adapters
Power Cord Selection
Contact PerkinElmer Technical Support (see page 7) to order
replacement power cords.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Fuses
Instrument Safety 4
WARNING
• Use only the power supply cord set provided with the Vectra
Polaris system. If the correct cord set for the location was not
provided, contact PerkinElmer Technical Support (see page 7)
for a replacement. Do not use power supply cords with
inadequate ratings.
• Use only a properly grounded power outlet when connecting the
system to power.
Contact PerkinElmer Technical Support (see page 7) to order
replacement fuses.
WARNING
The fuses in this instrument are only replaceable by trained
PerkinElmer personnel.
Cables and Adapters
Some cables and adapters supplied with the system have
proprietary specifications.
WARNING
Do not connect components supplied by PerkinElmer using
unqualified cables or adapters. Contact PerkinElmer technical
support (see page 7) to order replacement cables and adapters.
Mechanical Safety
WARNING
Instrument components may move during operation. Always keep
body parts, hair, jewelry, and clothing away from the instrument
during operation.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
WARNING
Procedures which could result in injury may be performed only by
operators who have been warned of the potential hazards and have
received adequate training in performing the procedures in the
safest possible manner.
Weight Warning
WARNING
LIFTING HAZARD. The Vectra Polaris instrument weighs 185 lbs.
(84 kg). Do not move the Vectra Polaris instrument. Installing,
servicing, and moving the Vectra Polaris instrument should be
performed only by qualified PerkinElmer service personnel. Contact
PerkinElmer technical support if help is required (see page 7).
Instrument Safety 5
Bright Light
WARNING
BRIGHT LIGHT HAZARD. The interior of the Vectra Polaris system
includes a barcode reader with a Class 2 LED Light. Do not look
into the bright light to avoid eye injury.
CAUTION - CLASS 2 LASER RADIATION WHEN OPEN
DO NOT STARE DIRECTLY INTO THE BEAM
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Copyright
Trademarks
Patents
Content
Preface 6
Preface
This manual is published by PerkinElmer, Inc., 68 Elm Street, Hopkinton, MA
01748 USA. Copyright 2017-2018, PerkinElmer, Inc., and its parent, affiliated, and
subsidiary companies. All rights reserved, including but not limited to those rights
to reproduce this publication or parts thereof. Reproduction of this publication or
parts thereof, or the products it describes, by any means or in any form is
expressly prohibited without the written permission of PerkinElmer.
PerkinElmer is a registered trademark of PerkinElmer, Inc. Microsoft and Windows
are either registered trademarks or trademarks of Microsoft Corporation in the
United States and/or other countries. All other trademarks are the property of their
respective owners.
US Patents 5,892,612; 5,953,087; 7,655,898; and patents pending.
Any errors or omission which may have occurred in this publication despite the
utmost care taken in its production will be corrected as soon as possible, but not
necessarily immediately upon detection. PerkinElmer provides this publication “As
Is” without warranty of any kind, either express or implied, including but not limited
to the implied warranties of merchantability or fitness for a particular purpose.
Some states or jurisdictions do not allow disclaimer of express or implied
warranties in certain transactions; therefore, this statement may not apply to you.
PerkinElmer reserves the right to revise this publication and to make changes from
time to time in the content hereof without obligation of PerkinElmer to notify any
person of such revision or changes. Further, PerkinElmer may make modifications
to the product described in this manual at any time without any obligation to notify
any person of such modifications.
Proper Equipment Operation
WARNINGS
• To reduce the risk of electric shock, do not remove the instrument panels. No
user serviceable parts are inside. Refer to qualified service personnel if help is
required.
• Use this product only in the manner described in this manual. If the equipment
is used in a manner not specified by the manufacturer, the protection provided
by the equipment may be impaired.
AVERTISSEMENTS
• Pour réduire le risque de choc électrique, ne pas retirer le couvercle. Ce
produit ne contient aucune pièce pouvant être réparée par l’utilisateur. Au
besoin, confier l’appareil à un réparateur qualifié.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Contact Us
Preface 7
• Ce produit ne doit être utilisé que comme décrit dans ce manuel. Si cet
appareil est utilisé d’une manière autre que celle spécifiée par le fabricant, la
protection fournie par l’appareil peut être entravée.
If you have a question about a product that is not answered in this manual or online
Help, or if you need assistance regarding this product, please contact the
PerkinElmer Technical Support Center from 8:00 A.M. to 8:00 P.M., Eastern Time,
Monday through Friday:
Phone: (US Toll Free): 800-762-4000
(Worldwide): +1 203-925-4602
Fax: +1 203-944-4904
Email: global.techsupport@perkinelmer.com
Internet: www.perkinelmer.com
Before you call, have the following information available for the technical
representative:
• Product serial number
• Software version (found by choosing About from the main Help menu)
• If applicable, the error number shown on the product’s LCD display, in the
software, or in the log file.
Product Service and Customer Support Plans
PerkinElmer offers a full range of services to ensure your success. From our
original factory warranty through a comprehensive line of customer support plans,
PerkinElmer offers you Field Service Engineers and in-house Specialists who are
dedicated to supporting your hardware, software, and application development
needs.
Phone: (US Toll Free): 800-762-4000
(Worldwide): +1 203-925-4602
Fax: +1 203-944-4904
Email: global.techsupport@perkinelmer.com
Our programs can include such useful services as:
• Preventive maintenance
• Diagnostic servicing performed on-site by PerkinElmer field service engineers
or remotely via Technical Support
• Validation performed on-site by PerkinElmer field service engineers
• Extended use of the PerkinElmer Technical Support Center
• Software updates
• Parts, labor, and travel expense coverage
• Other customized services upon request
Training For Your Product
Contact PerkinElmer for information about the availability of training courses for
your product:
Phone: (US Toll Free): 800-762-4000
(Worldwide): +1 203-925-4602
Fax: +1 203-944-4904
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
CE
KC
Preface 8
This device complies with all CE rules and requirements.
NOTE
Changes or modifications to this equipment not expressly approved by the party
responsible for compliance could void the user’s authority to operate the
equipment.
REMARQUE
Tout changement ou modification apporté à cet instrument non expressément
approuvé par l’entité responsable de la conformité peut annuler l’autorisation
d’opérer l’appareil accordée à l’utilisateur.
This device complies with MSIP (Ministry of Science, ICT, and Future Planning)
EMC Registration requirements. This instrument is registered as a Class A
instrument for business use only. Product seller and user should notice that this
equipment is not for household use.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Table of Symbols
Table 1 contains symbols that identify particularly important information and alert
you to the presence of hazards. These symbols may appear in this manual and/or
on the product it describes.
Preface 9
Table 1. Important Symbols
Symbol
Symbole
Description
Description
DANGER: An imminently hazardous situation, which, if not
avoided, will result in death or serious injury.
DANGER: Situation présentant un danger imminent qui, s’il
n’est pas éliminé, peut entraîner des blessures graves, voire la
mort.
WARNING: Caution. Refer to the User’s documentation. (ISO
7000-0434B)
AVERTISSEMENT: Attention. Se reporter à la documentation
de l’utilisateur.
NOTE: A cautionary statement; an operating tip or maintenance
suggestion; may result in instrument damage if not followed.
REMARQUE: Énoncé indiquant une précaution à prendre, un
conseil de fonctionnement ou une suggestion d’entretien; son
non-respect peut provoquer des dommages à l’instrument.
Hazardous voltage; risk of electric shock. (IEC 60417-6042)
Tension dangereuse; risque de blessure par électrocution.
Crush hazard. Risk of body parts, hair, jewelry, or clothing
getting caught in a moving part. (ISO 3864)
Danger d’écrasement. Faire attention que les parties
corporelles, les cheveux, les bijoux ou les vêtements ne soient
pas pris dans une pièce mobile.
Risk of fire. (ISO 3864)
Risque d’incendie.
Risk of explosion. (ISO 3864)
Risque d’explosion.
Lifting hazard. May result in injury. (ISO 3864)
Levage dangereux. Peut entraîner des blessures.
Protective ground symbol. (IEC 60417-5019)
Symbole de terre de protection.
Ground symbol. (IEC 60417-5017)
Symbole de terre.
Fuse. (IEC 60417-5016)
Fusible.
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Table 1. Important Symbols (Continued)
Preface 10
Symbol
Symbole
Description
Description
Alternating current. (IEC 60417-5032)
Courant alternatif.
On (power). |(IEC 60417-5007)
Marche (alimentation).
Off (power). (IEC 60417-5008)
Arrêt (alimentation).
CE compliance mark.
Marque de conformité CE.
Serial Number (ISO 7000-2498)
Numéro de série.
Date of Manufacture. (ISO 7000-2497)
Date de fabrication.
WEEE symbol (EN50419:2005). Do not dispose of as unsorted
municipal waste. See the PerkinElmer website
(www.perkinelmer.com) for more information.
Unified Korea Certification Mark (KC Mark)
Equipment
labels are color
coded:
Les étiquettes
de l’appareil
sont codées
couleur:
Signifies that the unit has passed safety tests for grounding,
power line transience, and current leakage.
Signifie que l’appareil a réussi les tests de sécurité pour la mise
à la terre, le courant transitoire de ligne d’alimentation et la
perte de courant.
Input.
Entrée.
Output.
Sortie.
Yel low Caution, risk of danger
Red Stop
Blue Mandatory action
Green Safe condition or information
Jaune Attention, danger potentiel
Rouge Arrêter
Bleu Intervention obligatoire
Ver t Condition sûre ou informations de sécurité
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
11
Table of Contents
Instrument Safety ................................................................................................... 2
Required Training................................................................................................2
Electrical Safety .................................................................................................. 3
Power Cord Selection ....................................................................................3
Fuses ............................................................................................................4
Cables and Adapters .....................................................................................4
Mechanical Safety...............................................................................................4
Weight Warning...................................................................................................5
Bright Light..........................................................................................................5
Preface .................................................................................................................... 6
Introduction........................................................................................................... 13
Intended Use..................................................................................................... 13
Principles of Operation...................................................................................... 14
Example Applications...................................................................................16
Theory of Imaging .............................................................................................16
Light ............................................................................................................ 17
Human Perception of Light Intensity and Color ............................................18
Light Absorbance and Reflection .................................................................19
Fluorescence ...............................................................................................20
Photobleaching............................................................................................ 21
Filter Sets for Conventional vs. Multispectral Imaging..................................22
Multispectral Analysis ..................................................................................24
Whole Slide Scanning .......................................................................................27
Multispectral Imaging ........................................................................................29
Hardware Reference ............................................................................................. 30
Front View......................................................................................................... 31
Top View........................................................................................................... 33
Right-Side View.................................................................................................34
Left-Side Connectors.........................................................................................35
Slide Carrier Hotel.............................................................................................36
Slide Carrier...................................................................................................... 38
Barcode Reader ................................................................................................39
System Computer and Monitor ..........................................................................40
System Computer Connectors ..................................................................... 40
Specifications....................................................................................................41
Hardware Operation.............................................................................................. 43
System Startup..................................................................................................44
Turn on the Vectra Polaris Instrument..........................................................44
Launch the Vectra Polaris Software.............................................................44
System Shutdown ............................................................................................. 45
Inspecting Slides and Slide Carriers..................................................................45
Inspect Slides .............................................................................................. 45
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12
Inspect Slide Carriers .................................................................................. 45
Loading Slides into the Slide Carriers................................................................ 46
Loading Slide Carriers into the Slide Carrier Hotel............................................. 47
Removing Slide Carriers from the Slide Carrier Hotel ........................................49
Removing Slides from the Slide Carriers ........................................................... 49
Software Operation............................................................................................... 50
Software Overview ............................................................................................ 51
Check Dashboard ........................................................................................ 52
Edit Protocol ................................................................................................52
Scan Slides .................................................................................................52
Launch Phenochart...................................................................................... 52
Gear Menu...................................................................................................53
System Dashboard............................................................................................ 54
Disk Space ..................................................................................................54
Brightfield References ................................................................................. 54
Fluorescence References ............................................................................56
Compensation Information...........................................................................57
Creating and Editing Protocols .......................................................................... 58
Studies ........................................................................................................ 58
Creating Protocols ....................................................................................... 59
Editing Protocols.......................................................................................... 60
Scanning Slides ................................................................................................78
The Carrier .................................................................................................. 78
Carrier Status ..............................................................................................78
Slide Status ................................................................................................. 80
Setting Up Scan Rules.................................................................................80
Maintenance.......................................................................................................... 89
Cleaning the Instrument Exterior .......................................................................89
Cleaning the Monitor ......................................................................................... 90
Cleaning the Power and Communication Ports.................................................. 90
Cleaning the Slide Carriers................................................................................90
Replacing the Fuses.......................................................................................... 90
Appendix A: PerkinElmer TIFF Specification ...................................................... 91
PerkinElmer Software License Agreement .......................................................... 97
Index.................................................................................................................... 107
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Introduction
This manual describes the use and functionality of the Vectra
PolarisAutomated Quantitative Pathology Imaging System. It
includes operating instructions, functional descriptions,
troubleshooting, illustrations, and other relevant information.
This section of the manual contains the following topics:
• Intended Use
• Principles of Operation
• Theory of Imaging
• Whole Slide Scanning
• Multispectral Imaging
Intended Use
The Vectra Polaris is a multimodal digital pathology instrument that
integrates both multispectral analysis and automated slide scanning
that allows researchers to visualize, analyze, quantify and
phenotype immune cells in situ in FFPE tissue sections and TMAs.
Introduction 13
NOTE
PerkinElmer's Vectra Polaris Quantitative Pathology Imaging
System is for research use only. Not for use in diagnostic
procedures.
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Principles of Operation
PerkinElmer’s Vectra Polaris is an automated imaging system for
performing whole slide scans of tissue sections and microarrays
(TMAs), and for acquiring multispectral (MSI) regions of interest.
The system has been optimized to image samples stained with
PerkinElmer’s Opal
reagent kits and is also compatible with typical brightfield staining
reagents.
The Vectra Polaris is configured to store and inventory up to 20
slide carriers, each holding up to 4 tissue slides, for a total of 80
slides.
multiplexed fluorescent immunohistochemistry
Introduction 14
Figure 1. The Vectra Polaris System
The Vectra Polaris has been designed to expand and support the
workflow defined in PerkinElmer’s Vectra® 3:
• True whole slide scanning of slides at 1.0 um/pix, 0.5 um/pix,
and 0.25um/pix
• Review and annotation of whole slides scans for MSI acquisition
• Acquisition of MSI regions
• Analysis of MSI regions including protein expression and
phenotyping
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Introduction 15
To implement the full capabilities of the instrument and workflow,
the Vectra Polaris system includes the following PerkinElmer
software:
• Vectra Polaris: Operator-centric software for performing whole
slide scans and acquiring MSI regions of interest. The Vectra
Polaris software runs on the workstation connected to the
Vectra Polaris instrument.
• Phenochart
: Whole-slide viewer and annotator of fluorescent
and brightfield scans acquired by the Vectra Polaris. Phenochart
allows the user to view the whole slide (zoom, pan, etc.), and
make decisions (annotations) on next steps for the sample.
Annotations in Phenochart are also used to record the workflow
actions for each slide scan. Annotations include reviewer
requested MSI fields; automated (inForm) field requests; and
reviewer edits, approvals, and rejections. The annotation file is
a fully auditable transaction log. Phenochart is freely distributed
and can be used by multiple users who wish to view or review
slide scans taken by the Vectra Polaris.
• inForm® Tissue Finder: Software typically used for the
analysis of MSI images. inForm supports features such as
tissue classification and training, cell phenotyping, protein
expression measurements, and data export. It can be run on the
Vectra Polaris computer and other Microsoft® Windows 10
computers. Additional inForm software “seats” beyond the two
seats that come with each Vectra Polaris system are available
for purchase.
Vectra Polaris workflows range from simple two-step procedures
(e.g., acquire whole slide scan and review) to automated acquisition
of regions of interest selected by the user or the Vectra Polaris
itself. An example fluorescence workflow might include the following
steps:
1 Stain tissue with PerkinElmer Opal fluorescent IHC reagents.
2 Acquire whole slide fluorescent imagery using the Vectra
Polaris.
3 Review the whole slide imagery with Phenochart and annotate
regions of interest for MSI analysis.
4 Acquire the MSI regions with Vectra Polaris.
5 Use inForm to phenotype cells and measure protein expression
levels in the acquired MSI regions.
For example applications for the Vectra Polaris, see Example
Applications.
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Example Applications
Some examples of Vectra Polaris applications include:
• Whole slide scanning and multispectral interrogation of tissue
samples and microarrays stained with Opal reagent kits
• Whole slide scanning of tissue samples stained with H&E and
conventional IHC stains
• Phenotypic analysis and protein expression of immune and
cancer cells in the context of the tumor microenvironment.
Theory of Imaging
This section introduces some important concepts used by
PerkinElmer’s Vectra Polaris imaging systems, including:
• Light
• Human Perception of Light Intensity and Color
• Light Absorbance and Reflection
Introduction 16
• Fluorescence
• Photobleaching
• Filter Sets for Conventional vs. Multispectral Imaging
• Multispectral Analysis
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Light
Introduction 17
For purposes of this discussion, light refers to the part of the
electromagnetic spectrum that can be seen by the human eye and
the nearby ultraviolet and infrared wavelengths. While the physical
description of light can be highly complex, we will focus on these
wavelengths of light and how they interact with physical and
biological materials.
Figure 2. The Electromagnetic Spectrum
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Human Perception of Light Intensity and Color
Response to Illumination
The human eye is a highly adaptive light detector. It is significantly
more sensitive in low light than in bright light. When light levels
change, it takes some time for the eye to fully adjust. This is the
reason people need to “dark adapt” in a darkened room before they
are able to observe weak fluorescence through a microscope.
Humans can see both in very dark and very bright settings.
Because the eye is so adaptable to various lighting conditions,
humans are unable to quantify absolute levels of light. In any given
situation, the eye has a limited ability to discriminate levels of
illumination. US Department of Defense research indicates that
most humans can only distinguish approximately 30 to 35 levels of
gray, ranging from black to white.
The eye's response to illumination is not linear. It more closely
approximates a logarithmic function. The result is that the human
eye cannot see small proportional changes in brightness.
Introduction 18
Contrasting the eye with conventional microscope imagery, any
sensor that has 8-bit resolution can detect 256 levels of gray. As the
number of bits of resolution increase, the number of gray levels also
increases. A 12-bit sensor yields 4096 levels of gray. Digital
electronic sensors are linear in response to light levels.
Ability to Distinguish Colors
While the eye is relatively poor at discriminating intensity, it is very
good at distinguishing colors. Most individuals are able to
discriminate thousands of colors. However, no two individuals see a
given color in exactly the same way. The eye contains three
different types of color sensors, also known as cone cells. While the
arrangement of cone cells is generally standard from person to
person, the ratio of each type of cone cell varies, as does their
actual physical arrangement within the eye. These minor variations
(along with the brain's interpretation of the color) lead to the
differences in perceived color between individuals.
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Light Absorbance and Reflection
We perceive objects based on the way they transmit, absorb, and/or
reflect light.
Absorbance and reflection work in tandem. Absorbance refers to
the wavelengths of light that are 'taken in by' the objects. This
means that an object that we perceive as red has absorbed all
visible wavelengths of light except red. The red wavelengths are
reflected back to the eye of the observer.
Transmittance refers to light emitting objects such as light sources,
and fluorescing or phosphorescing objects. An object we would
perceive as red in transmission is one that transmits primarily red
wavelengths, while absorbing or reflecting other wavelengths.
In brightfield light microscopy we observe light that passes through
a specimen. Except for a few pigments and inclusions, biological
specimens are essentially invisible. To impart contrast, we employ
some absorbing dye, or specific optical arrangement. It is this need
for contrast that led to the initial development of biological stains
and stain protocols and subsequently to phase contrast and other
optical contrast enhancing techniques.
Introduction 19
Optical Density (OD) is used to measure the interaction of light with
absorbing materials. The science of absorbing spectroscopy is
based on the Beer-Lambert law. When absorbing images collected
in brightfield are converted to OD images, the information contained
in each pixel is quantitative, as to the amount of absorbing material
present. PerkinElmer's brightfield multispectral imagery is
automatically converted to optical density at acquisition time,
enabling quantitative analysis.
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Fluorescence
Introduction 20
Many biological and natural materials give off light of a particular
color when exposed to light of another color. This property is a type
of luminescence. There are two types of luminescence:
• Fluorescence refers to luminescence that occurs when the light
is emitted rapidly after illumination (around one-millionth of a
second).
• If the light emission takes longer than one-millionth of a second,
the luminescence is called phosphorescence.
Materials that exhibit fluorescence have proven extremely useful as
labels or indicators in many biological systems.
Fluorescence light emission is different than light absorption. Each
fluorescent molecule generates light. Fluorescent light can be
measured quantitatively because it does not interact with other
materials. While it would seem that fluorescence could be
measured more accurately than absorbed light, there are a number
of factors that complicate such measurements. For example, light
scatters, it is affected by the local environment (such as pH), and
the measurements can be affected by surrounding molecules.
Stokes Shift
When you excite a specimen with a particular (shorter) wavelength
(such as blue), the specimen then shines in a different (longer)
wavelength (such as red, orange, or yellow). The difference
between the wavelength of the (shorter) exciting light and the
wavelength of the (longer) emitted light is called the Stokes Shift,
which is based on Stokes Law.
The wavelength or color you use to excite the specimen (i.e. the
'excitation light') and the color the specimen glows (i.e. the
'emission light') depend on the dye involved. For any given
fluorescence dye, there will be a range of excitation wavelengths
that will excite fluorescence. This range of excitation wavelengths is
known as the absorption spectrum. Each dye also emits across a
range of wavelengths, known as the emission spectrum. The figure
below contains an example of excitation and emission spectra,
showing Stokes Shift and the overlap of the spectra.
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Introduction 21
Figure 3. Stokes Shift
In addition, many biological materials are naturally fluorescent. This
is known as autofluorescence. In particular, many vitamins, some
hormones, and a variety of biological enzymes and structural
proteins are naturally fluorescent. These materials often fluoresce
strongly enough to interfere with specific fluorescence labeling
studies.
Photobleaching
Because dyes can be damaged by intense light, reducing the
emission signal (‘photobleaching’), it is important to limit the time
they are exposed to excitation light or to bright light during routine
handling. Usually, blue or UV light is the most damaging. The
Vectra Polaris uses an electronically-gated excitation source
synchronized with its camera so the sample is only exposed to light
while the camera is taking an image. Also, the Polaris front door is
made of a translucent plastic that absorbs harsh blue and UV light.
When using Vectra Polaris, avoid spending a long time in the
Protocol Exposures Editor while fixed on any one spot of the
sample, since it takes a live image stream.
These steps enable repeated measurements with minimal effect on
the sample.
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Introduction 22
Filter Sets for Conventional vs. Multispectral Imaging
Filters used for conventional fluorescence imaging are often
designed so they only transmit a very narrow range of wavelengths
of light. Limiting the measurement to wavelength bands where the
dye is inherently most responsive helps distinguish the desired dye
from other dyes or background signals in the sample. In this way, it
is possible to image several dyes, provided that their spectra are
separated rather than overlapping. Based on the properties of
common dyes, this puts an upper limit of ~4 on how many dyes can
be imaged in any one sample.
An alternative approach is to image the sample multispectrally. In
this case, a broad emission filter is used, and a tunable filter is
engaged in the imaging path. The instrument takes pictures at
several wavelengths within the emission band, so maps out the full
shape of the dye response. This enables analysis software to
identify what dye(s) are present, in what amounts, in each pixel, by
spectral decomposition (“unmixing”). It also enables identifying, and
removing, contributions from autofluorescence.
These two approaches are illustrated in Figure 4 and Figure 5.
Figure 4. Narrowband excitation (solid
line) and emission (dashed line) filter
for conventional imaging.
Ideally, the excitation filter would match the excitation maximum of
the fluorescence label being used, and the emission filter would
include the emission maximum. In practical terms, the filter maxima
may be slightly different from the ideal case, due to limitations of
filter manufacturing and because for many dyes the Stokes shift is
small, so the maxima are quite close to one another.
Figure 5. Narrowband excitation (solid
line) and long-pass emission filter
(dashed line) for multispectral imaging.
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Introduction 23
To image multiple dyes conventionally, one selects dyes that have
very distinct excitation and/or emission response, and selects filters
that are narrow enough to predominantly transmit the signal of only
one dye at a time (Figure 6 and Figure 7).
Figure 6. Imaging four dyes by
conventional fluorescence methods.
This shows the emission spectra for four dyes (DAPI, FITC, Cy3,
and Cy5.5) that are spectrally fairly distinct, and can be separated
by conventional imaging. This works because there is only a little
overlap between adjacent dyes. Excitation filters and dye response
are omitted for clarity, but are similarly separable.
This approach breaks down when more dyes are present, or when it
is important to account for the effects of autofluorescence. Figure 7
shows the spectra of 7 dyes, and with this many dyes there is no
way to isolate their signals using conventional imaging techniques,
or to account for autofluorescence.
Figure 7. Seven dyes is too many to
image by conventional methods.
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Multispectral Analysis
The Vectra Polaris imaging system offers a unique solution to the
problem of separating the signals from highly multiplexed samples.
Multispectral analysis is based on the fact that all fluorescent
materials produce a unique spectral emission. If you excite a
material and examine the emitted fluorescence over a range of
wavelengths, the resulting emission intensities can generate an
“emission spectrum”. This spectrum is different for each specific
fluorescent material. For many fluorescent labels of biological
interest, the emission spectra overlap, and may be further obscured
by autofluorescence from the specimen. Multispectral imaging
provides a way to distinguish between many overlapping emission
spectra within the same area, overcoming the limitations of
conventional filter-based imaging. With the additional information
provided by the LCTF during MSI imaging the system can
distinguish between dyes with fully overlapping spectra within a
single channel (Figure 8).
Introduction 24
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Introduction 25
Figure 8. Unmixed multispectral image of human breast cancer
tissue stained against CD4 (Green), CD20 (Red), CD8 (Yellow),
FoxP3 (Orange), CD68 (Purple), Cytokeratin (Light Blue), and
DAPI (Dark Blue) using Opal reagents.
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Introduction 26
In general, multispectral analysis generates the spectral curves for
the various fluorescent dyes or materials in a specimen. In addition,
it generates a spectral curve for the autofluorescence that almost
always is present to some degree. Using spectral analysis
algorithms, the contribution of the individual fluorescence spectra
are separated. The result is a set of images representing each
spectrum that contributes to the final image 90 (Figure 9).
Figure 9. Removing auto-fluorescence with multispectral
imaging. A) Conventional fluorescence image of tonsil tissue
stained against CD4 with Opal 520 dye. B) Overlay of all 35
layers of a multispectral image acquired using five filter cubes.
C) Emission spectra of pure autofluorescence (black line), pure
Opal 520 (green line), and the mixture observed in 'B' (gray
dashed line). The spectral references are used to 'unmix' the
contribution of autofluorescence and Opal 520 at each pixel. D)
Image of unmixed Opal 520 signal extracted from 'B' with > 10fold higher signal-to-background compared to 'A' because the
autofluorescence contribution has been separated into the
component image in 'E'.
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Whole Slide Scanning
Vectra Polaris scans slides using the following process:
1 Color Overview:
Vectra Polaris takes a low power color overview of all four slides
in each carrier, including the label for each slide. This initial step
is performed regardless of whether you are using a
fluorescence or brightfield protocol and is used to identify the
presence of slides in the carrier and capture their labels.
2 Coverslip Finding:
For each slide, Vectra Polaris will then find the coverslip using
this overview scan. The coverslip defines the potential scan
area.
Introduction 27
For fluorescence protocols, you can further restrict this area by
making a closed loop with a red, green, or blue Sharpie
marker. If closed-loop markup is present, the system will only
scan within the loop. This is useful if your tissue is faintly
stained, punctate, or if you have highly fluorescing non-tissue
material (PAP pen, for example.)
For brightfield protocols, the coverslip will define the potential
scan area. Closed-loop markup is not available for brightfield
scans.
3 Slide Height Finding:
Vectra Polaris engages specialized height-sensing optics to
measure the top of the coverslip at up to 9 locations. This gives
an initial focus estimate based on the expected coverslip
thickness.
4 Fluorescence Overview (Fluorescence protocols only):
If you are using a fluorescence protocol for this slide, Vectra
Polaris will take another overview within the coverslip (or
closed-loop markup), this time in fluorescence.
5 Tissue Finding:
®
Using the corresponding overview, Vectra Polaris will
automatically detect the sample on the slide. The resulting area
will be scanned. If requested within the protocol, Vectra Polaris
will scan the entire area within the coverslip (or closed-loop
markup).
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Introduction 28
6 Focus Finding:
Vectra Polaris will measure focus at multiple points on the tissue
to determine best focus. It uses the sample map from the
previous step to choose the measurement grid, and continues
until the grid is fully measured.
If the measured tissue height is irregular, Vectra Polaris
increases the grid density and takes more readings until it finds
the readings are regular at the newly finer scale.
All focus measurements include a dust-rejection algorithm, and
an overall consistency check is applied as well, to further reduce
the likelihood of dust-induced focus errors.
7 Scanning:
Vectra Polaris then scans the slide.
Brightfield scans are conventional color scans that have been
color and background corrected.
Fluorescence scans are multi-layered, with one layer for each
filter you chose. To avoid photobleaching, the system uses a
pulsed LED so the sample is only exposed to light during the
time that the camera is taking a picture.
When scans are complete, they can be opened in Phenochart,
the whole slide viewing application.
NOTES
Some dyes narrowly express in a single filter. Other dyes may
express in multiple filters and may appear in more than one layer in
a Vectra Polaris fluorescence scan. For example, Opal 570 will
have signal in both Cy3 and TexasRed filters. If your sample is
highly multiplexed, multiple dyes may appear in the same channel.
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Multispectral Imaging
Once Vectra Polaris has completed a whole slide scan of the tissue,
individual regions can be selected for multispectral imaging.
Multispectral imagery is acquired using the following process:
1 Selection of Multispectral Regions:
Regions for multispectral imaging are selected on a previously
scanned slide. Using Phenochart, you can select individual
fields or regions of interest. If desired, you can also select fields
using a trained inForm algorithm. See the Phenochart
documentation for more information.
When you configure that slide for MSI acquisition, Vectra Polaris
will perform the following actions:
2 Color Overview:
Vectra Polaris takes a low power color overview of all four slides
in each carrier, including the label for each slide. This initial step
is performed regardless of whether you are using a
fluorescence or brightfield protocol and is used to identify the
presence of slides in the carrier.
Introduction 29
3 Slide Registration:
Using the above overview along with the slide’s original
overview, Vectra Polaris accounts for any shift or rotation of the
slide to ensure that the multispectral region locations are
accurate. The slide edges are used to account for any rotation
or horizontal shift. The coverslip edges to account for vertical
shifting.
4 Slide Height Finding:
Vectra Polaris engages specialized height-sensing optics to
measure the top of the coverslip at up to 9 locations. This gives
an initial focus estimate based on the expected coverslip
thickness.
5 Acquisition of Multispectral Regions:
Vectra Polaris will then travel to each multispectral region site,
autofocus, correct for exposure if requested, and acquire the
multispectral image.
Multispectral imagery can then be viewed and analyzed in
inForm.
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Hardware Reference 30
Hardware Reference
This section identifies and describes the Vectra Polaris system
hardware. It also lists the Vectra Polaris technical specifications.
WARNING
Lifting Hazard. Do not move the Vectra Polaris instrument.
Installing, servicing, and moving the Vectra Polaris instrument
should be performed only by qualified PerkinElmer service
personnel. Contact PerkinElmer technical support if help is required
(see page 7).
This section contains the following information:
• Front View hardware components
• Top Vie w hardware components
• Right-Side View hardware components
• Left-Side Connectors
• Slide Carrier Hotel
• Slide Carrier
• Barcode Reader
• System Computer and Monitor
• Specifications
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Front View
Door
Slide
Carrier
Hotel
Sensor
Status
Progress
Display
Hardware Reference 31
Figure 10 shows the front view of the Vectra Polaris instrument with
the system door open. The parts identified are described below the
figure.
Figure 10. Vectra Polaris Front View - System Door Open
Status Progress Display
Each light represents an individual slide and indicates the progress
of the slide currently being scanned.
Slide Carrier Hotel
Houses up to 20 slide carriers. See Slide Carrier Hotel on page 36
for more detailed information.
Door Sensor
Opens or closes the System Door when a hand is placed in front of
the sensor.
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Hardware Reference 32
Slide
Carrier
Status
Lights
System
Door
Closed
Figure 11 shows the front view of the Vectra Polaris instrument with
the system door closed. The parts identified are described below
the figure.
Figure 11. Vectra Polaris Front View - System Door Closed
Slide Carrier Status Lights
Each slide carrier status light represents an individual slide carrier
and indicates the processing status of each Slide Carrier. See
Tab le 1 for the color codes of the slide carrier status lights.
System Door
When closed, covers the Slide Carrier Hotel and slide processing
can occur. When open, reveals the Slide Carrier Hotel.
WARNING
Do not operate the system if there has been a malfunction of the
system door or slide loading components. Contact PerkinElmer
technical support if help is required (see page 7).
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Top View
Cover
Handle
Hardware Reference 33
Figure 12 shows the top view of the Vectra Polaris instrument. The
part identified is described below the figure.
Figure 12. Vectra Polaris Top View
Cover Handle - Used by only service personnel to remove
instrument covers during service and installation.
WARNING
DO NOT remove instrument covers. There are no user serviceable
parts inside. The covers are intended to be removed by qualified
service personnel only; they are not intended to be removed during
operation or for maintenance by users. Contact PerkinElmer
Technical Support if help is required (see page 7).
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Right-Side View
System Power Switch
Figure 13 shows the right-side of the Vectra Polaris instrument. The
part identified is described below the figure.
Hardware Reference 34
Figure 13. Vectra Polaris Right-Side View
System Power Switch
Turns the Vectra Polaris instrument ON (I) or OFF(O).
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Left-Side Connectors
Power
Connector
USB 2.0 Connector
USB 3.0
HUB
Connector
Figure 14 identifies the connectors on the bottom left-side of the
Vectra Polaris instrument. The connectors identified are described
below the figure.
Figure 14. Vectra Polaris Left-Side Connectors
Power Connector - Connects to the power cord to provide power to
the Vectra Polaris instrument.
WARNING
Hardware Reference 35
• Use only the power supply cord set provided with the Vectra
Polaris system. If the correct cord set for the location was not
provided, contact PerkinElmer Technical Support (see page 7)
for a replacement.
• Use only a properly grounded power outlet when connecting the
system to power.
USB 2.0 Connector- Connects a USB 2.0 cable to a USB 2.0 port
on the System Computer Connectors.
USB 3.0 Hub Connector- Connects a USB 3.0 HUB to a USB 3.0
port on the System Computer Connectors.
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Slide Carrier Hotel
Slide Carrier
Hotel Slot 1
Slide Carrier
Status Lights
The Slide Carrier Hotel houses up to 20 Slide Carriers and is visible
when the System Door opens. Figure 15 shows a closeup of the
Slide Carrier Hotel and identifies its components. The components
are described below the figure.
WARNING
Do not operate the system if there has been a malfunction of the
system door or slide loading components. Contact PerkinElmer
technical support if help is required (see page 7).
Hardware Reference 36
Figure 15. Slide Carrier Hotel
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Hardware Reference 37
Slide Carrier Status Lights
Indicates the processing status of each slide carrier. Tab le 1 lists
the color codes associated with the slide carrier status lights.
Table 1. Slide Carrier Status Lights - Color Codes
Color Status
None Slide carrier hotel is empty.
White Initial state of slot after slide carrier is inserted
and no rules have been applied via software.
Solid Red Hotel slot is malfunctioning.
Blinking Red Slide carrier is not completely inserted.
Blue Awaiting Processing.
Blinking Blue Prioritized and awaiting processing.
Yellow Processing instructions are either incomplete
or invalid.
Blinking Green Processing.
Solid Green Processing Complete.
Orange Processing complete but a processing error
occurred.
Hotel Slot
Each hotel slot holds one Slide Carrier. The hotel slots are
numbered 1-20, starting at the top of the hotel.
Slide Carrier
Houses up to four slides. See Slide Carrier for more detailed
information.
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Slide Carrier
Slide Position 1
Slide
Carrier
Spring-Loaded Tab
Insert IndicatorSlide Position 4
Tab C o ver
Handle
Slide Carriers are an integral component of Vectra Polaris. The
Slide Carrier holds up to four microscope slides and helps protect
slides from damage. The microscope slide positions s are labeled 1
to 4 and correspond to slide identification in the Vectra Polaris
software.
Figure 16 shows a closeup of a Slide Carrier and identifies its
components. The identified components are described below.
Hardware Reference 38
Figure 16. Vectra Polaris Slide Carrier
Slide Position
Each slide position is populated with one slide. Slide position 1 is
on the far left; slide position 4 is on the far right.
Slide
Each microscopic slide is manually loaded into the slide positions.
The table below identifies the details of the slide format.
Slide Type Width Height Thickness
Metric 25.0 ± 1.0mm 75.0 ± 1.0mm 1.00 ± 0.10mm
English 25.4mm 76.2mm 1.00 ± 0.10mm
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Insert Indicator
A Perkin Elmer icon that indicates the side of the slide carrier
to be inserted into a Hotel Slot.
Tab Cov er
Holds the spring-loaded tabs in place.
Spring-Loaded Tab
When a slide is inserted into a slide slot, it is gently placed up
against a spring-loaded tab.
Carrier Handle
The side of the slide carrier to hold when inserting and removing a
slide carrier from a Hotel Slot. A unique number is printed on top of
the handle for slide carrier identification purposes.
Barcode Reader
Hardware Reference 39
The Vectra Polaris is equipped with an internal Barcode Reader.
The Barcode Reader reads the barcode on the slide labels.
WARNING
BRIGHT LIGHT HAZARD. The barcode reader includes a Class 2
LED Light. Do not look into the bright light to avoid eye injury.
CAUTION - CLASS 2 LASER RADIATION WHEN OPEN
DO NOT STARE DIRECTLY INTO THE BEAM
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System Computer and Monitor
USB 2.0 Port
USB 3.0 Port
The Vectra Polaris system includes a widescreen monitor and a
computer pre-installed with Vectra Polaris, Phenochart, and inForm
software. A wireless keyboard and mouse are also included. See
System Computer Connectors for a description of the computer
connectors.
System Computer Connectors
Figure 17 shows the Vectra Polaris computer connectors. The
connectors identified are described below the figure.
Hardware Reference 40
Figure 17. System Computer Connectors
USB 2.0 Port - Connects a USB 2.0 cable to the USB 2.0 connector
on the instrument’s Left-Side Connectors.
USB 3.0 Port - Connects a USB 3.0 HUB to the USB 3.0 HUB
connector on the instrument’s Left-Side Connectors.
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Specifications
This section lists the technical specifications for the Vectra Polaris
instrument. Technical specifications are subject to change without
notice.
NOTE
Vectra Polaris is for research use only. Not for use in diagnostic
procedures.
System
Size (H x W x L) 28” (72 cm) x 30” (77 cm) x 27.25” (69.2 5cm)
Weight 185 lbs. (84 kg)
Spectral Range 440 nm to 720 nm
Objectives 4x, 10x, and 20x
Hardware Reference 41
Pixel Resolution 0.25 um, 0.5 um, or 1.0 um
File Format PerkinElmer proprietary .qptiff format for
whole slide scans, .im3 file format for
multispectral data; 24- bit Windows-compatible
bitmap for RGB/Mono imagery
Operating
System
RAM 16GB
Microsoft® Windows 10, 64-bit
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Hardware Reference 42
Environmental
Operating Temperature 59°F to 83°F (18°C to 28°C)
Operating Humidity 10% - 50% non-condensing
Storage Temperature 59°F to 86°F (15°C to 30°C)
Storage Humidity 0% - 80% relative humidity, non-
condensing
Altitude Up to 2000m (6560ft.)
Shipping Temperature
14°F to 113°F (-10°C to +45°C)
(up to 72 hours max)
Pollution Degree 2
Indoor Use Only
Electrical
One properly grounded AC power outlet for the computer, monitor,
and instrument must be located within 6 feet (1.8 m) of the location.
Input Voltage 100 to 230 VAC (±10%), 500 W, 50/60 Hz
System does not have transient overvoltage
protection.
Computer Interface USB 2.0, USB 3.0
Fuse 4A Littelfuse® 250V, 5mm x20mm
Barcode Reader Light Source (White laser diode)
Wavelength 400 nm to 750
Power Output 70mAmps
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Hardware Operation 43
Hardware Operation
This section describes how to operate the Vectra Polaris hardware.
It is important to read and understand Instrument Safety on page 2
before using the system. If you are not familiar with the Vectra
Polaris system hardware, Hardware Reference on page 30 provides
a description of each hardware component in the Vectra Polaris
system.
This section includes the following procedures:
• System Startup on page 44
• System Shutdown on page 45
• Inspecting Slides and Slide Carriers on page 45
• Loading Slides into the Slide Carriers on page 46
• Loading Slide Carriers into the Slide Carrier Hotel on page 47
• Removing Slide Carriers from the Slide Carrier Hotel
• Removing Slides from the Slide Carriers
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
System Startup
This section describes the procedures needed to start the Vectra
Polaris system:
• Turn on the Vectra Polaris Instrument
• Launch the Vectra Polaris Software
• Inspect Slides
• Inspect Slide Carriers
Turn on the Vectra Polaris Instrument
To start the Vectra Polaris instrument:
1 If necessary, plug the Vectra Polaris power cord from the Left-
Side Connectors into an appropriate power outlet.
WARNING
Hardware Operation 44
• Use only the power supply cord set provided with the Vectra
Polaris system. If the correct cord set for the location was not
provided, contact PerkinElmer Technical Support (see page 7)
for a replacement. Do not use power supply cords with
inadequate ratings.
• Use only a properly grounded power outlet when connecting the
system to power.
• The wall outlet or the power cable connector on the left side of
the instrument should be accessible after the system's
installation, to enable trained service personnel to safely
disconnect power from the system during servicing.
2 Turn on the computer and allow Windows 10 to start.
3 Switch the System Power Switch to the ON (I) position. The
Status Progress Display flash green to indicate a quiescent
state. The Door Sensor is responsive and functional.
Launch the Vectra Polaris Software
To launch the Vectra Polaris software:
1 Click the Vectra Polaris icon on the Windows 10 desktop.
2 The Vectra Polaris homepage opens (see page 50).
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Hardware Operation 45
System Shutdown
This section describes how to shut down the Vectra Polaris system.
To shutdown the Vectra Polaris system
1 Exit the Vectra Polaris software. If open, the System Door
closes.
2 Select Shut down from the Windows Start Menu to power down
the computer.
3 Switch the System Power Switch on the right side of the
instrument to the OFF (O) position. The status lights turn off and
the system shuts down.
Inspecting Slides and Slide Carriers
Before inserting slides into slide carriers, both should be inspected
for potential defects.
This section describes the steps needed to properly Inspect Slides
and Inspect Slide Carriers.
Inspect Slides
•Verify the Slides meet the required formats and dimensions.
• Do not use broken or damaged slides, or slides with broken or
damaged coverslips.
• Use only slides that are free of debris, fingerprints, and dust.
Inspect Slide Carriers
•Verify the Slide Carrier Tab Cover is secure.
• Do not use slide carriers that are warped or bent.
• If any sticky residue is on the carrier handle or outer surfaces,
clean before use.
NOTE
See page 90 for slide carrier cleaning instructions.
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Hardware Operation 46
Loading Slides into the Slide Carriers
After the Slides and Slide Carriers have been successfully
inspected, slides can be loaded into the Slide Carriers.
To load a slide into a slide carrier:
1 Place the slide carrier onto a flat surface.
2 Hold the microscope slide by the label end with the coverslip
side up.
3 Gently push the opposite end of the slide into the desired Slide
Position until the slide is up against the Spring-Loaded Tab.
Figure 18. Insert Slide into Slide Carrier
4 Gently lay the label end of the slide completely into the slide
position. The tab should push the slide against the opposite wall
of the slide position.
5 Load up to four slides into each slide carrier. Slides need not be
contiguous.
6 Click Enter Slide IDs (see page 80) in the Vectra Polaris
software. The Enter Slide IDs dialog opens.
7 Type the applicable slide IDs into the text boxes and click the
OK button.
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Hardware Operation 47
8 To load additional slides into another slide carrier, repeat steps
1 to 7 above. Slide carriers may be stacked on top of one
another for easy handling and storage (see Figure 19).
Figure 19. Slide Carrier Stack
Loading Slide Carriers into the Slide Carrier Hotel
The Slide Carrier Hotel stores Slide Carriers before and after
microscope slide scanning. The slide carrier hotel can store up to
20 slide carriers for a total of 80 slides. The Hotel Slots are
numbered 1-20, starting at the top of the hotel.
NOTES
Slide carriers can be loaded into the slide carrier hotel before or
after launching the software.
To load a slide carrier into the slide carrier hotel:
1 Inspect the slide carrier (see page 45).
2 Verify the Vectra Polaris System is on (see page 44).
3 If the System Door is closed, place your hand in front of the
Door Sensor to open it. The system doors slides to the left to
reveal the slide carrier hotel.
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Hardware Operation 48
Carrier
Insert
Indicator
Handle
4 Hold the slide carrier by the Carrier Handle with the slide labels
facing upwards.
5 Gently insert the end of the slide carrier with the Insert Indicator
() into a Hotel Slot.
Figure 20. Ends of a Slide Carrier
6 When the Slide Carrier is halfway into the Hotel Slot, the carrier
engages a roller and creates a small amount of friction. Push
the carrier further into the slot until it is fully seated.
Figure 21. Push the Slide Carrier until Fully Seated
• If the carrier is inserted correctly, a click noise indicates that
the carrier has been seated correctly into the slot. The Slide
Carrier Status Light (page 37) next to associated Hotel Slot
turns white.
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Hardware Operation 49
• If the carrier is not inserted correctly, the Slide Carrier Status
Light next to associated the Hotel Slot blinks red until the
slide carrier is removed from the slot or is positioned
correctly.
7 Continue to load up to 20 slide carriers into their slots.
8 When done, place your hand in front of the Door Sensor to close
System Door.
Removing Slide Carriers from the Slide Carrier Hotel
When slide scanning is complete, the Slide Carriers can be
removed from the Slide Carrier Hotel.
To remove a slide carrier from the hotel:
1 Pull the Carrier Handle until the slide carrier is free from its
Hotel Slot. The Slide Carrier Status Lights next to the Hotel Slot
turns off, indicating no slide carrier is inside the Hotel Slot.
2 Lay the slide carrier onto a flat surface.
Removing Slides from the Slide Carriers
After Slide Carriers are removed from the Slide Carrier Hotel, the
microscopic slides can be removed from the Slide Carriers.
To remove a slide from a slide carrier:
1 Gently push against the slide label end of the microscope slide
using your index finger, compressing the tab on the far end of
the slide.
2 Using the same finger, lift the label end of the microscope slide
from its slot.
3 Grab the label end of the slide with your thumb and index finger.
4 Remove the remaining end of the slide from the slide carrier.
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Software Operation 50
Software Operation
This section describes how to operate the Vectra Polaris software.
This section includes the following topics:
• Software Overview
• System Dashboard
• Creating and Editing Protocols
• Scanning Slides
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Software Overview
The Home Page links to the pages needed to maintain and run Vectra Polaris.
Software Operation 51
The Home Page buttons link to the following pages:
Check Dashboard - Check your remaining disk space and acquire references.
Edit Protocol - Create or edit brightfield and fluorescent protocols and studies.
Scan Slides - Select the rules to scan your slides and scan your slides based on
those rules.
Launch Phenochart - Launch the Phenochart viewing software.
You can also get to those pages by using the Gear Menu (in the upper right):
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Additionally, you can use the Gear Menu to check your version number or perform
other functions, including launching online help.
Below is an overview of each page.
Check Dashboard
Use the Dashboard to check the status of your system prior to imaging. It includes
the following:
• Disk space - checks the available space on the disk drive where images will be
stored.
• Acquire References - views and acquires brightfield references and
fluorescent references.
For detailed information on how to use the System Dashboard, see System
Dashboard.
Edit Protocol
Use the Edit Protocol page to define protocols and take snapshots.
Software Operation 52
A protocol describes how a sample is to be imaged, including the imaging mode
(brightfield or fluorescence), and the spatial resolution (magnification) for the
whole slide scan and for multispectral regions of interest. For fluorescent imaging,
it also describes the exposure times and what bands to use for focusing and
imaging.
For detailed information on how to create and edit protocols and studies, see
Creating and Editing Protocols.
Scan Slides
Use the Scan Slides page declare how slides should be imaged. Slide scanning
can be started and stopped from this page.
For detailed information on how to perform whole slide scans, see Scanning
Slides.
Launch Phenochart
Use this button to launch the Phenochart program.
For detailed information on how to use Phenochart, see the Phenochart User's
Manual or online Help from within the Phenochart program.
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Gear Menu
Use the Gear menu (in the upper right hand corner of the Home Page) to:
• Link to the following pages:
• Obtain license information and upgrade your license
• Find out What's New in the current installed version
• Launch the online Help system
• Open the Preferences dialog
• Open the Vectra Polaris software Log for reference or troubleshooting
• Open the About window to view the current software version
• View contact information for PerkinElmer technical support
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— Check Dashboard
— Edit Protocol
— Scan Slides
— Launch Phenochart
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System Dashboard
Use the System Dashboard to:
• See the available Disk Space
• View and acquire Brightfield References
• View and acquire Fluorescence References
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Disk Space
The disk space bar shows where your data is stored and how much space is
available on that drive.
Brightfield References
Click View to see reference imagery and information for the label image,
brightfield overview, color image and multispectral image.
• Export For Diagnostics will save this image to aid in technical support.
• Show raw images will display the images without scaling, which is useful for
technical support.
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Click Acquire to take new brightfield references. Use the stage control to:
• Move around to a clean area in the live view (no coverslip lines, tissue, or
label) using the stage navigation tool.
• Click within the slide to change positions. You can refine the position by using
arrow keys for small movements, and control+arrow for slightly larger
movements.
• Click OK to take references.
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Fluorescence References
Click View to see reference imagery and information for the overview, 20x and
40x resolution references for each filter.
• Export For Diagnostics will save this image to aid in technical support.
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Click Acquire to take new fluorescent references. We recommend you take
references for all filters at once, but you can take references for a single filter if
necessary. Acquiring references for all filters can take over 30 minutes, and the
system can be left unattended during this time.
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Compensation Information
Brightfield
Acquired images of a sample are normalized (divided) by the reference images on
a pixel-by-pixel, wavelength-by-wavelength basis. This yields the sample
transmission T, which is in the range 0 - 1. This transmission (T) is then mapped:
• For multispectral images, the Optical Density is calculated - this is based on
Log10(T).
• For simple color images, like a whole slide scan, the transmission is mapped
from 0 - 255.
Fluorescence Protocols
Acquired images are normalized by a shading pattern derived from the reference
images. The shading pattern, which is the reference image divided by the mean
intensity in the center, is applied on a pixel-by-pixel basis, with one pattern per
epi-filter. Here, center means the central 1/16
shape of the shading pattern varies per instrument, the overall effect is to increase
the signal near the image edges, and to do little or nothing to the signal from the
center of the image.
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th
of the image area. While the exact
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Creating and Editing Protocols
This section contains the following topics:
• Studies
• Creating Protocols
• Editing Protocols
Studies
Vectra Polaris stores scan data by ‘Study’.
A study is a group of slides that belong together. This could be an experimental
study (e.g. Ki67 markers in breast cancer tissue), all slides from one source, or
other groupings. Each study contains one or more slides. Each slide may be
scanned more than once, if needed.
• The default location for a study is D:\Data\VectraPolaris\[Study] (where [Study]
is the name of the study).
• Whole-slide scans and supporting imagery acquired from specific slides are
saved to slide-specific subfolders in the main study folder. See the section on
Scanning Slides for more details about imagery.
• MSI Fields acquired from a particular slide scan will be stored in an MSI folder
within the scan folder.
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A protocol defines the set of rules to be used during whole slide and multispectral
region acquisition, including imaging mode, pixel resolution, filter cubes, exposure
times, and other parameters.
• Protocols have the file extension “.ppr” and are saved in
D:\Data\VectraPolaris\[Study].
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Creating Protocols
Before you create a protocol, it is helpful to know how the slide was stained.
• Brightfield protocols are used to acquire imagery from slides stained with H&E
or conventional chromogenic IHC methods.
• Fluorescence protocols are used with PerkinElmer Opal and other fluorescent
dyes.
To create a new protocol:
•Select Edit Protocol from the Vectra Polaris Home Page.
• Click New and the following window will appear:
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• Entera Protocol name.
•Select Brightfield or Fluorescence under Imaging Mode.
• Select a previously created Study or create a new Study.
— To select a previously created study, click on the study in the Available
Studies list. This will highlight the study.
— To create a new study, enter the Study Name in the text box and click
the Create Study button. The new study will be added to the Available
Studies list where you can select the study.
• Click the Create button to create the protocol in the selected study.
To load an existing protocol, click Load and select the protocol from the study
folder.
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Editing Protocols
After you have created your brightfield or fluorescence protocol and assigned it to
a study, use the Edit Protocol screen to add specific details to the protocol. The
next sections are organized by two different types of protocols: brightfield and
fluorescence. The sections appear as follows:
• Brightfield Protocols
• Brightfield Snapshots
• Fluorescence Protocols
• Fluorescence Exposures
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Brightfield Protocols
After you have created your brightfield protocol and assigned it to a study, the Edit
Protocol screen (for brightfield protocols) will appear:
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Under Whole Slide Scan, choose the Pixel Resolution that you want to use to
image the whole slide. You can choose 1 um per pixel (nominally 10x), 0.5 um per
pixel (nominally 20x), or 0.25 um per pixel (nominally 40x).
If you plan to take Multispectral Regions, choose the Pixel Resolution that you
want to use for imaging. Again, available resolutions are 1 um per pixel (nominally
10x), 0.5 um per pixel (nominally 20x), or 0.25 um per pixel (nominally 40x).
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Advanced Settings
There may be situations when you need to fine-tune how Vectra Polaris scans the
slides. Click the Advanced Settings button to adjust the following functions:
• Vectra Polaris is designed for use with standard #1.5 coverslips, where the
distance between the tissue and the top of the sample is approximately
170 microns. While this is the optimal setup, Vectra Polaris can handle other
scenarios.
— If your lab uses thinner coverslips (#0), select “100 um (#0 or similar)”
for a Coverslip Thickness.
— If your lab uses thicker coverslips, or has a significant amount of
mounting media between the sample and coverslip, select the option
that best matches your slides.
NOTE
Vectra Polaris is capable of handling slides with up to 420 microns of combined
coverslip or mounting medium above the sample, though imagery at these
extremes may be sub-optimal.
• If Vectra Polaris is having difficulty finding your entire tissue sample, check the
Scan within the entire coverslip region checkbox. This will increase scan
time and file size, but will enable you to complete scanning of difficult samples.
• Vectra Polaris has a specialized algorithm for finding TMA samples. If your
samples are TMAs, select Sample is a TMA.
• By default, brightfield whole slide scans in Vectra Polaris are .jpg compressed
in order to save disk space. You can adjust the image quality as needed;
higher quality will result in larger files. If you would like use lossless
compression instead, select LZW rather than JPEG.
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• If you would like Vectra Polaris to decode the barcode on your slide label and
save it as part of the scan file, select the type of barcode you are using. By
default, Vectra Polaris will not attempt to decode a barcode unless you
specifically select this option.
— If you have only 1D barcodes, select All 1D barcodes (autodetect
type).
— If your lab uses a variety of 2D barcodes, select All 2D barcodes
(autodetect type).
NOTE
If your lab only uses a specific type of 2D barcodes, you can decrease the
decoding time by selecting that specific type. Vectra Polaris supports decoding of
Data Matrix, PDF 417, QR Code, Aztec, and Maxicode.
• If your lab is using a combination of 1D and 2D barcodes, select All barcodes
(autodetect type). This option will increase scan time and file size, and should
only be used if necessary.
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Click Save to save the protocol. The study you previously chose will be
automatically selected. You can change the protocol name or study if needed.
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Brightfield Snapshots
Once you have selected your scan and MSI resolutions, you can take example
snapshots of your slides.
Click Take Snapshots to load the Brightfield Snapshots editor.
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If a carrier is not already on stage, click Load Carrier to select a carrier that
contains the slides you wish to image.
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Use the navigation tool to select an area on the slide. Click on the slide to change
positions.
For easier navigation, click Take Overview to acquire imagery of all slides in your
carrier. You can also select that option in the Preferences dialog in the Gear
Menu. Vectra Polaris will then automatically acquire overview imagery whenever a
carrier is loaded.
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Click on the tissue to change positions.
You can refine the position by using arrow keys (for small movements) or
control+arrow (for slightly larger movements).
Click Autofocus or use the Stage Height slider to bring the live view into focus.
You can click on a feature within the live view to center on that location or click
and drag within the live view to change locations.
To take a snapshot of your current live view, select either the whole slide or the
MSI regions radio button, pick a base file name, and click Snap.
You can navigate to new places, focus as needed, and take as many snapshots as
you want. Snapshots will be numbered incrementally.
Click Back when you are ready to return to the protocol editor.
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Fluorescence Protocols
When creating a fluorescence protocol, you will need slides that include positive
expression in all markers of interest in order to set suitable exposure times.
After you have created your fluorescence protocol and assigned it to a study, the
Edit Protocol screen (for fluorescence protocols) will appear:
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Under Overview Scan Rules, select the desired filter that will be used to help find
tissue on the slide. This will typically be your DAPI counterstain. If there is no
counterstain, then choose a filter that aligns with the expression of your most
common stain or auto-fluorescence.
Under Whole Slide Scan, choose the Pixel Resolution to be used while imaging
the slide. Options include:
• 1.0 um per pixel (nominally 10x)
• 0.5 um per pixel, (nominally 20x)
• 0.5 um per pixel, (nominally 20x, binned from a 40x acquisition)
• 0.25 um per pixel (nominally 40x).
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NOTE
Each time you change the pixel resolution, you will need to reset the exposure
times.
• You can auto-update your exposures. This is recommended as a first step to
give a close approximation of your exposure times. We then recommend you
use the exposures editor to fine-tune your exposures.
• You can manually update your exposures. In this case, the current exposures
remain unchanged and you must use the exposures editor to obtain valid
exposures.
• You can cancel. This will leave the pixel resolution unchanged.
If you plan to take Multispectral Regions, choose the Pixel Resolution that you
want to use to image the MSI regions. Available resolutions are 10x, 20x, 20x
binned, or 40x.
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The Edit Filters and Bands button allows you to choose which filters you use to
take your whole slide scan and which bands you use for multispectral imaging.
Imagery for each filter or band will be taken in the order it is shown.
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Whole Slide Scan
By default, Vectra Polaris will take a whole slide scan with all five filters. If you
would like to save disk space, choose only the filters where fluorescent signals
are expressing. You can remove a filter by clicking on the red 'x' next to that filter.
Click Add Filter to add an additional filter. You can change the colors associated
with each filter; these are the colors that Phenochart will use to display the whole
slide scan.
Multispectral Imaging
In contrast, in multispectral imaging, we strongly recommend that you maintain
the default band configuration. Non-signal bands are integral to multispectral stain
extraction and unmixing.
Advanced Settings
There may be situations when you need to fine-tune how Vectra Polaris scans the
tissue. Click the Advanced Settings button to adjust the following functions:
• Vectra Polaris is designed for use with standard #1.5 coverslips, where the
distance between the tissue and the top of the sample is approximately
170 microns. While this is the optimal setup, Vectra Polaris can handle other
scenarios.
— If your lab uses thinner coverslips (#0), select “100 um (#0 or similar)”
for a Coverslip Thickness.
— If your lab uses thicker coverslips, or has a significant amount of
mounting media between the sample and coverslip, select the option
that best matches your slides.
NOTE
Vectra Polaris is capable of handling slides with up to 420 microns of combined
coverslip or mounting medium above the sample, though imagery at these
extremes may be sub-optimal.
• You can specify which filter will be used for setting focus during whole slide
scans and for MSI imaging. Choose the filter where your counterstain
expresses. This is normally DAPI.
• Saturation protection prevents you from overexposure when acquiring
multispectral imagery. We highly discourage deactivating the Use Saturation
Protection setting.
• If Vectra Polaris is having difficulty finding your entire tissue sample, check the
'Scan within the entire coverslip region' checkbox. This will increase scan time
and file size, but will enable you to complete scanning of difficult samples.
NOTE
If there is Sharpie® markup on your fluorescent slide, Vectra Polaris will use that
as the region instead of the coverslip.
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• Vectra Polaris has a specialized algorithm for finding TMA samples. If your
samples are TMAs, select Sample is a TMA.
• If you would like Vectra Polaris to decode the barcode on your slide label and
save it as part of the scan file, select the type of barcode you are using. By
default, Vectra Polaris will not attempt to decode a barcode unless you
specifically select this option.
— If you have only 1D barcodes, select All 1D barcodes (autodetect type).
— If your lab uses a variety of 2D barcodes, select All 2D barcodes
(autodetect type).
NOTE
If your lab only uses a specific type of 2D barcodes, you can decrease the
decoding time by selecting that specific type. Vectra Polaris supports decoding of
Data Matrix, PDF 417, QR Code, Aztec, and Maxicode.
— If your lab is using a combination of 1D and 2D barcodes, select All
barcodes (autodetect type). This option will increase scan time and
file size, and should only be used if necessary.
Once you have selected your settings, use the Exposures Editor to set exposures
for your protocol.
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Filter Cube Recommendations when using PerkinElmer Opal Reagents
If you are using your Vectra Polaris system with PerkinElmer's Opal multiplex IHC
detection kits or with individual Opal fluorophores, please refer to the table below
for exposure time recommendations for each filter cube.
Filter
DAPI FITC Cy3 Texas Red Cy5
DAPI
Opal 520
Opal 540
Opal 570
Opal 620
Opal 650
Opal 670
Opal 690
Key
:
• Colors are signal bands: autoexpose or manually set the desired exposure per
sample
• Gray is non-signal band: manually set to 20ms exposure for MSI
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Fluorescence Exposures
Once you have selected your scan and MSI resolutions, filters, and bands, you
need to set exposures. Click Edit Exposures in the Fluorescence Protocol Editor
to set exposure times.
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If a carrier is not already on stage, click Load Carrier to select a carrier that
contains representative slides. Select your carrier from the list and load it to the
stage.
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Use the navigation tool to select a slide. You will see a live view of your first filter
for your imaging rules. In most cases, this will be DAPI. The highlighted entry in
the table identifies which filter or band is currently shown.
For easier navigation, click Take Overview to acquire digitally enhanced imagery
of all slides in your carrier. You can also select that option in the Preferences
dialog in the Gear Menu. Vectra Polaris will then automatically acquire overview
imagery whenever a carrier is loaded.
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Click on the tissue to change positions.
You can refine the position by using arrow keys (for small movements) or
control+arrow (for slightly larger movements).
You can click on a feature within the live view to center on that location, or click
and drag within the live view to change locations.
Click Autofocus or use the Stage Height slider to bring the live view into focus.
Click Autoexpose to have the system find the best exposure for that filter/band.
• After auto-exposing, you may want to auto-focus and auto-expose again to
refine your focus and exposure estimates.
• You can override the auto-exposure value by typing a value in the highlighted
cell. Values must be between 0.1 and 2000 ms.
• If you would like to turn off the fluorescence illumination and live view, click
Close Shutter. You will need to re-open the shutter to see a live view and set
exposures.
If you plan to use this protocol for multispectral region acquisition, you should set
the MSI exposures at this time. In most cases, you can use the same area of the
slide to set your MSI exposures. Click the MSI regions radio button to set
exposures for MSI regions. Focus and click Autoexpose.
• As with whole slide scan exposures, after auto-exposing, you may want to
auto-focus and auto-expose again to refine your focus and exposure estimates.
• You can override the auto-exposure value by typing a value in the highlighted
cell. Values must be between 0.1 and 2000 ms.
• If you would like to turn off the fluorescence illumination and live view, click
Close Shutter. You will need to re-open the shutter to see a live view and set
exposures.
Repeat the steps above for all filters and bands in your protocol. You may need to
change locations and/or slides to find the best signals for setting your exposures.
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NOTE
if there is an N/A in the table, that filter or band is not in use, so no exposure is
needed.
Snapshots
When you have set your exposures, you can take snapshots to see sample
imagery of how your exposures perform on a given area of the slide. Select either
the Whole Slide or the MSI Regions radio button, pick a base file name, and click
Snap. You can navigate to new places and take as many snapshots as you want.
They will be numbered incrementally.
Click Back when you are ready to return to the protocol editor.
NOTE
The exposures you just set are only valid for the resolutions you picked. If you
change your scan or MSI resolution, you will need to revisit the exposures editor
to update your exposures.
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Scanning Slides
The section describes the procedures involved when scanning slides:
• The Carrier
• Carrier Status
• Slide Status
• Setting Up Scan Rules
The Carrier
Slides are loaded into Vectra Polaris via carriers, which are stacked into the Slide
Carrier Hotel.
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The hotel can hold up to 20 carriers, and each carrier can hold up to 4 slides. This
means Vectra Polaris can be loaded with up to 80 slides at any given time. Those
80 slides can all be scanned based on the same set of rules, or you can tailor your
scan rules to each slide and/or each carrier as needed.
Carrier Status
Within the software, each hotel slot is identified as ‘Slot N,’ where N is 1-20
running top to bottom. When a slide carrier is loaded into the hotel, an LED light
changes color to identify the status of that carrier.
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Carrier status icon
Slide status icon
Slide ID
The User Interface will reflect the current state of each slot in the hotel. If there is
no carrier present, you will see:
Each carrier present in the hotel will be represented as follows:
Each carrier is represented by a carrier status icon, four slide status icons, and
four slide IDs.
The carrier status icon corresponds to the hotel LED for that particular carrier.
Each possible status is represented by a different color.
When you insert the carrier into the hotel, a white icon indicates that the carrier
is present but has no rules to use for slide scanning. If the icon is blinking red ,
the carrier has not been completely inserted. Try pushing it in further.
Once the carrier has been inserted in the hotel, you can define rules for your scan.
The icon will change to blue when there is at least one slide ready to be
scanned. Carriers that have been prioritized will be blinking blue .
If no slides on the carrier can be scanned with the current rules, the icon will
change to yellow . This carrier will be ignored until the problem is resolved.
When scanning has started, the icon for the carrier on stage will be blinking green
. When the carrier is returned to the hotel, the icon will either be green (for a
successful scan) or orange (if there was a problem with the scan).
If the icon is solid red , that hotel slot’s sensor is malfunctioning. Do not use this
slot.
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Slide Status
Each carrier also has a status icon for the 4 slides it contains.
means the slide has complete rules and is ready to be scanned.
(blinking) means the slide is currently being scanned.
means the slide was scanned successfully.
Other options include , which means the slide has rules, but they cannot be
used. (They may be incomplete, in which case you will need to edit the rules for
that slide). indicates that the slide failed to scan correctly.
Finally, will indicate that the slide is being ignored. This is useful if the carrier
has fewer than four slides or if you don’t want to process that slide at this time.
Setting Up Scan Rules
Scan rules must be defined for each slide you would like to process. Scan rules
are complete when a study, protocol, task, and slide ID have been assigned.
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Slide IDs
You must manually enter the ID for each slide in the carrier. You can enter IDs
before the carrier is placed into the hotel, or you can enter IDs after the carrier has
been placed into the hotel.
If you would like to enter the IDs while the carrier is still in your hand, click Enter
Slide IDs.
It will bring up this dialog:
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You can then enter between 1-4 slide IDs for your slides and click OK.
Immediately put the carrier in the hotel, and Vectra Polaris will assign the slide IDs
you entered to that carrier.
If your carrier is already present in the hotel, you can edit the carrier to add the
slide IDs along with the other scanning rules.
Editing a Single Slide
To edit the scanning rules for a single slide, click the status icon for that slide. It
will bring up this editor:
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For this slide, enter:
• A task: Choose Scan or Take M SI (Note: Select Ignore if you do not wish to
process this particular slide).
• A study: Select a study.
• A protocol: Select any protocol you have saved within the study.
• The Slide ID source: Select manual entry and add your slide ID, if necessary.
Check Prioritize the containing carrier if you want this slide’s carrier to be
scanned at the front of the line.
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Editing All Slides Within a Single Carrier
To edit the scanning rules for a single carrier, click the status icon for that carrier.
It will bring up this editor:
For each slide, you can enter:
• A task: Choose Scan, Take MSI or Ignore. (Note: select Ignore if there is no
slide in that particular slot).
• A study: Select a study.
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• A protocol: Select any protocol you have saved within the study.
• The Slide ID source: Select manual entry and add your slide ID, if necessary.
Check Prioritize this carrier if you want this carrier to be scanned at the front of
the line.
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Editing Multiple Carriers
To edit multiple carriers, Click Configure Tasks.
Select the carriers you want to edit. Multiple selection is available.
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You will also need to select how you want to process your slides.
To process all slides using the same rules, choose Using the same rules for all
slides and click Next… The Edit Slides dialog will appear:
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• Select your task, study, and protocol.
• Select manual entry for your slide ID source.
• Check Prioritize the containing carriers if you want the selected carriers to
be scanned at the front of the line.
NOTE
You will not be able to edit the slide ID, because you are applying rules across
multiple slides. If you need to enter slide IDs, you can edit the individual carriers.
To use different processing rules for each carrier position, choose Using different
rules for each carrier slot and click Next… For each position in the carrier,
select the task, study, and protocol. Select manual entry for your slide ID source,
and prioritize if desired. Again, you will not be able to edit the slide ID, because
you are applying rules across multiple slides.
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Saving and Restoring Scan Setup
After setting up all the details for your slides, you may want to save your work for
later use. This may be useful if the scanning process is interrupted before the task
is finished or if you need to return later to rescan certain slides.
You can save your slide IDs, studies, protocols, and tasks by clicking the Save
Setup button. Vectra Polaris will also prompt you to save before scanning (when
you click Scan). At this point, you can enter a name and some notes about this
scan setup.
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Saving a setup is not required, but it can help you locate the information later if
needed. By default, this information will be deleted after 30 days, but you can
override the default and opt to keep the information until you manually delete it.
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To restore an earlier scan setup, click Load Setup.
All available setups will be shown. You can filter by date to show setups only from
a specific time period. Click on the headers to sort by date, name, notes, studies,
etc. (Note: You can also delete entries that are no longer needed by selecting
those items and clicking Delete). To use a setup, select it from the dialog and click
Load.
Once you have selected a setup, you need to configure the carriers in Vectra
Polaris to match the information in the setup.
If the carriers and slides are the same location as when you saved the setup, you
only need to click OK, because everything is in the right place.
In other cases (for example: when you need to put your slides back in), this dialog
will help you figure out where your slides belong.
1 When scan information is available and a carrier is present in the hotel, the
system will show the slide IDs (slot 1 in the image below)
2 When scan information is available but there is no carrier present in the hotel
slot, the slot will be labeled as Missing in red (see slot 5 below). The slide IDs
for that carrier are listed, so you can locate the correct slides, place them in
the carrier, and insert the carrier into the hotel.
3 When no information is available for a carrier that is present in the hotel, the
system will show italicized Slide N text for the slide IDs (slots 7, 9, etc). Slides
in these carriers were not present in the original setup.
For every item that the scan setup had - meaning, anywhere the system has
information on a carrier that should be present (items 1 and 2) - you need to
specify what to do before the carrier can be loaded. There are 3 options:
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1 Use as saved: The system will load all slide information including the scan
task.
2 Ignore task: The system will load all slide information except for the scan
task. The scan task will be set to ignore. This is useful if you are loading the
setup so you can rescan a few select items, or if you know that you're going to
be changing the task from how it was originally saved.
3 Don't restore: No information for this carrier will be reloaded. This is useful if
you do not have or do not need the carrier.
The red text at the bottom of the screen will list any issues that need to be fixed
before scanning.
Click OK when you have finalized your selections. The scan slides editor will be
cleared and replaced with your selected scan information.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Scanning
Software Operation 88
The Scan button will enable when at least one carrier is ready to be scanned.
Click Scan to start scanning. If any carriers have been prioritized, these carriers
will be processed first.
Vectra Polaris will report scanning progress for each slide on the progress dialog
and on the front panel LEDs.
You click Pause Scanning to remove completed carriers and add new carriers to
be scanned. When you pause a scan, Vectra Polaris will return the current carrier
to the hotel. It will then be safe to open the door. You can remove carriers that are
completed and replace them with unscanned carriers. Edit those carriers to set
them up for scanning, and reprioritize carriers if needed. When you’re ready, click
Scan to resume scanning.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Maintenance
This section includes procedures that are to be performed
periodically, either to make the Vectra Polaris run better, or to
protect its components from damage.
• Cleaning the Instrument Exterior on page 89
• Cleaning the Monitor on page 90
• Cleaning the Power and Communication Ports on page 90
• Cleaning the Slide Carriers on page 90
• Replacing the Fuses on page 90
Cleaning the Instrument Exterior
Clean the Vectra Polaris instrument exterior as necessary.
The non-electrical exterior parts of the Vectra Polaris can be wiped
down with a soft cloth using standard laboratory grade cleaning
solutions including:
Maintenance 89
• 70% ethanol
• 10% bleach
• Clidox®
• Sporicidin®
WARNING
DO NOT spray cleaning solutions directly onto the Vectra Polaris
instrument. Sprays and liquids that come into contact with the
Vectra Polaris instrument may result in damage to the system or
electric shock. Always spray the cleaning solution onto a cloth and
then wipe the instrument exterior with the cloth.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Maintenance 90
Cleaning the Monitor
Clean the monitor as necessary with a soft, lint-free cloth. If
needed, dampen the cloth with water or an eyeglass cleaner.
WARNING
DO NOT spray cleaning solutions directly onto the monitor screen.
Sprays and liquids that come into contact with the screen may
result in damage to the system or electric shock. Always spray the
cleaning solution onto a cloth and then wipe the screens with the
cloth.
Cleaning the Power and Communication Ports
Dust can be removed from the power and communication ports with
a gentle air stream from compressed air cans. DO NOT shake the
can before use. Hold can at a distance from the ports to prevent
condensation.
WARNING
Turn off the electrical power to the Vectra Polaris system by
shutting down the system (see page 45) before cleaning any part of
the instrument where electrical or fiber optic cables make
connections.
Cleaning the Slide Carriers
It is important to keep carriers free from sticky debris caused by
slide labels, tape, or mounting media.
Slide carriers can be cleaned using soap and water with a soft
cloth, or using an ultrasonic bath.
Replacing the Fuses
For blown fuses and to order replacement fuses, contact
PerkinElmer Technical Support (see page 7).
WARNING
Electric Shock Hazard. DO NOT attempt to replace the fuses. Only
qualified PerkinElmer service personnel can replace the fuses.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Appendix A: PerkinElmer TIFF Specification 91
Appendix A: PerkinElmer TIFF Specification
This describes a TIFF format that PerkinElmer uses for its tissue images. The imagery may
be a simple RGB image, a set of unmixed multispectral image (MSI) components, or a
whole-slide scan. In the latter case, it may be a brightfield (BF) color RGB image or a
multiband fluorescence (FL) image.
The goal is to use the same syntax and metadata for all these kinds of images, and
minimize the semantic distinctions where possible. Specifically, the unmixed MSI images
can be considered to be an idealized multiband FL image where signal corresponds to the
presence of a stain or fluorescent dye in the sample.
Metadata
Metadata is contained in two locations: within standard TIFF tags as listed in the table
below, and within the ImageDescription string, using a set of XML tags described below.
These are provided for each image (IFD) in the file, and describe that image rather than the
baseline image or the scan as a whole. The ScanProfile tag is only provided on the first,
baseline image as it may be large.
Data format
The files are TIFF or BigTIFF images, depending on image size, with multiple images per
file.
For images larger than about 2K x 2K pixels, tiled format is used, and the image is provided
in several resolutions (pyramidal tiled images). Tile size is 512 x 512 pixels. Images smaller
than 2K x 2K use stripped format.
The highest resolution (baseline) image(s) appear first in the file. For each resolution there
are N baseline images where N depends on the contents. For BF images, N=1 and each
image is an RGB image. For FL images or unmixed component images, N = number of
bands, which is usually > 1, and each image is a grayscale image.
A thumbnail RGB image is provided, and this is a good image to use as an icon in graphical
image lists. This comes after the baseline images, meaning it is the second image in BF
(RGB) images, and the (N+1)st image for FL images or unmixed component images.
Next come the reduced-resolution images (if present). The pyramid contains enough levels
that the image size is no larger than 2K x 2K at the coarsest resolution.
For whole-slide scans, there are two more non-tiled images after these: an optional RGB
image of the label, and a macro (low-resolution) RGB image of the whole slide.
Overall, the arrangement is:
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Appendix A: PerkinElmer TIFF Specification 92
Table A-1. Images
Description RGB/mono Title/Strip Resolution Notes
Baseline image Varies Varies Full Tiled if > 2K x 2K
RGB for BF, else mono
More full-resolution
Mono Varies Full If N > 1
images
Thumbnail RGB Stripped ~500 x 500
Half-resolution
Varies Varies Half Only if baseline is tiled
images
Quarter, eighth, etc. Varies Varies Quarter,
eighth, etc.
Macro (overview)
RGB Stripped ~2000 x 4000 Required for whole-
image of whole slide
Continues until 2K x 2K
or smaller
slide scans Optional
for simple RGB and
MSI components
Label image RGB Stripped ~500 x 500 Optional, whole-slide
scans
Detection
Readers can recognize PerkinElmer tissue images via the contents of the “Software” TIFF
tag (see below). The file suffix is .qptiff for whole slide scans.
Metadata
Metadata is contained in two locations: within standard TIFF tags as listed in the table
below, and within the ImageDescription string, using a set of XML tags described below.
These are provided for each image (IFD) in the file, and describe that image rather than the
baseline image or the scan as a whole. The ScanProfile tag is only provided on the first,
baseline image as it may be large.
Table A-2. TIFF tags
TIFF Tag Optional Description of contents
Software Starts with “PerkinElmer-QPI”
ImageDescription Further metadata in XML format (see next section)
ImageWidth Width of the image in pixels
ImageLength Height of the image in pixels
ResolutionUnit Unit used for resolution and position (see below)
XResolution Pixel X resolution (see below)
YResolution Pixel Y resolution (see below)
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Appendix A: PerkinElmer TIFF Specification 93
Table A-2. TIFF tags
TIFF Tag Optional Description of contents
XPosition Y Sample X location in ResolutionUnits. This is
ULHC location except for Macro image which
reports its image center.
YPosition Y Sample Y location in ResolutionUnits. This is
ULHC location except for Macro image which
reports its image center.
SampleFormat Integer (1) for BF, FL; or float (3) for unmixed MSI
images
SMinSampleValue Minimum signal value in the image
SMaxSampleValue Maximum signal value in the image
BitsPerSample 8 (FL); 8, 8, 8 (RGB); or 32 (unmixed component)
SamplesPerPixel 1 (FL or unmixed component) or 3 (RGB)
NewSubfileType 0 for full-resolution images, 1 for reduced res
images
TileWidth Y Tile width (512) if tiled format is used
TileLength Y Tile height (512) if tiled format is used
TileOffsets Y List of tile offsets, if tiled format is used
TileByteCounts Y Size of each (compressed) tile, if tiled format is
used
StripOffsets Y List of strip offsets, if tiled format is not used
RowsPerStrip Y Number of rows per strip, if tiled format is not used
StripByteCounts Y Size of each (compressed) strip, if tiled format is
not used
PlanarConfiguration 1 (chunky) for RGB images, 2 (planar) otherwise
PhotometricInterpretation 2 (RGB) for RGB images, 1 (BlackIsZero)
otherwise
DateTime Acquisition time
Compression May be None, CCITT Group 3, PackBits, LZW, or
JPEG
JPEG fields Y JPEG fields are defined when JPEG compression
is used
ResolutionUnit, XResolution and YResolution are required fields in a valid TIFF file. When
the true resolution of the image is known, ResolutionUnit will be 3 (cm) and XResolution
and YResolution will be pixels/cm. When the true resolution is not known, ResolutionUnit
will be 2 (inch) and XResolution and YResolution will be 96 (pixels/inch). PKI TIFF pixels
are always square so XResolution and YResolution will always have the same value.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
Appendix A: PerkinElmer TIFF Specification 94
The TIFF spec is not explicit about the data type and value for SMinSampleValue and
SMaxSampleValue; the PerkinElmer writer uses the same data type as the image pixels
(byte or float).
Image Description contents
The ImageDescription tag contains a string in XML format. The string contains a top-level <
PerkinElmer-QPI-ImageDescription> element. Nested within this element are child
elements with the tag names and values as listed in the table below. Elements appear in the
order listed. Values are stored as text content of the element. Elements are required unless
otherwise specified. See the example below.
Table A-3. Image Description tags
Tag Optional Contents
DescriptionVersion Version of the image description field itself, a
single number. This document describes version
2 of the field.
AcquisitionSoftware Software used to acquire the image.
Identifier GUID in string format. This is an identifier for the
image file itself.
SlideID Y ID of the slide that this image was taken from.
Barcode Y Barcode text of the slide this image was taken
from.
ComputerName Y Name of the computer on which the slide was
scanned.
ImageType A string identifying the type of image within the
file (Table S-1), with the following values:
• FullResolution
• ReducedResolution
• Thumbnail
•Overview
• Label
IsUnmixedComponent “True” for unmixed multispectral images,
otherwise “False”.
ExposureTime Exposure time as an integer number of
microseconds. For unmixed images, this is the
exposure time for the dominant wavelength band
for the component (FL); or the brightest
wavelength in the cube (BF).
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Appendix A: PerkinElmer TIFF Specification 95
Table A-3. Image Description tags(Continued)
Tag Optional Contents
SignalUnits A byte wwww tttt where the tttt nibble indicates
the signal unit type from the following:
0–raw counts
1 –normalized (counts/second/gain/full-
scale/binning)
4 –OD (optical density)
5 –dark-corrected counts
and the wwww nibble indicates how the signal is
weighted across the spectral bands (or colors):
0 –average across all bands
2 –total summed signal across all bands
4 –peak signal in highest-valued band
Thus, for example, a value of 68 (hex 44)
encodes OD units with peak-signal weighting.
Name Y Band (component) name for FL or unmixed MSI
images, not present for RGB images.
Color Y Color to use when rendering this band, as
decimal r,g,b byte triplet, r FL or unmixed MSI
images. Not present for RGB images.
Responsivity Y Instrument responsivity, if available, for FL whole-
slide and unmixed images. See below for details.
Objective Y Objective name, if known, otherwise not present.
ScanProfile Y Element containing scan and/or and unmix
parameters. It is valid XML whose contents are
opaque to most readers. It is only provided on the
first (baseline) image, and is omitted from all
other IFDs.
ValidationCode Used for internal data integrity checks – readers
can ignore this.
For whole slide images (BF and FL), SignalUnits will be 64 (hex 40) (raw counts, peak
signal). For unmixed images, SignalUnits will reflect the unmix settings.
Instrument Responsivity
The <Responsivity> tag is a container for a list of normalized instrument response values.
This tag is present for whole-slide FL images from Vectra Polaris and unmixed FL MSI
images originating from Vectra 3 and Vectra Polaris.
For whole-slide images, the <Responsivity> tag will contain one <Filter> tag. The < Filter>
tag contents will be different for each image within the TIFF file, reflecting the filter used to
take the image.
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Appendix A: PerkinElmer TIFF Specification 96
For unmixed component images, the <Responsivity> tag will contain one <Band> tag for
each band in the original IM3 file; typically there will be five <Band> tags. The same
<Band> tags are repeated for each unmixed component image.
The overview, thumbnail and label images do not have <Responsivity> tags.
Each <Filter> or <Band> tag describes the instrument responsivity for acquisitions using
that filter or band. The contents of the <Filter> and <Band> tags are described below.
Table A-4. Contents of Filter and Band tags
Tag Contents
Name The name of the filter (whole-slide image) or MSI band (unmixed image).
Response The instrument response to the reference artifact, normalized for exposure.
This is raw counts / (2
bit depth
× exposure time × gain × binning area), where
bit depth is the bit depth of the imagery, exposure time is in seconds, gain is
the gain setting of the camera, and binning area is 1 for 1×1, 4 for 2×2, etc.
Date The date and time of the reference image in UTC, ISO 8601 format.
FilterID Detailed description of the acquisition filter.
Sample ImageDescription
Sample ImageDescription for the DAPI band of a FL whole-slide image, containing a single
<Filter> tag:
<?xml version="1.0" encoding="utf-8"?>
< PerkinElmer-QPI-ImageDescription >
<DescriptionVersion>1</DescriptionVersion>
<AcquisitionSoftware>VectraScan 1.0.0</AcquisitionSoftware>
<ImageType>FullResolution</ImageType>
<Identifier>AABED946-BB58-44FB-95B3-48E177E3BB83</Identifier>
<IsUnmixedComponent>False</IsUnmixedComponent>
<ExposureTime>50</ExposureTime>
<SignalUnits>64</SignalUnits>
<Name>DAPI</Name>
<Color>0,0,255</Color>
<Responsivity>
<Filter>
<Name>DAPI</Name>
<Response>30.7</Response>
<Date>2015-10-22T13:10:18.0618849Z</Date>
<FilterID>Semrock:FF02-409/LP-25 Emission / Semrock:FF01-377/50-25
Excitation</FilterID>
</Filter>
</Responsivity>
<Objective>4x</Objective>
<ScanProfile><!-- this will be a serialized scan protocol. It is valid XML but
otherwise opaque -->
</ScanProfile>
<ValidationCode>4281ff86778db65892c05151d5de738d</ValidationCode>
</ PerkinElmer-QPI-ImageDescription >
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
PerkinElmer Software License Agreement 97
PerkinElmer Software License Agreement
***IMPORTANT INFORMATION-PLEASE READ CAREFULLY***
The following is an agreement (the “Agreement”) between you and PerkinElmer, Inc., 68 Elm St., Hopkinton, MA 01748 (“PKI”) for software
known as Research Use Only Vectra Polaris, including any updates, upgrades or enhancement provided by PKI and any and all accompanying
documentation (collectively, the “Software”).
PKI IS WILLING TO LICENSE THE SOFTWARE TO YOU ONLY UPON THE CONDITION THAT YOU ACCEPT ALL OF THE TERMS
CONTAINED IN THIS AGREEMENT. PLEASE READ THE TERMS CAREFULLY BEFORE USING THE SOFTWARE, AS INSTALLING
AND/OR USING THE SOFTWARE WILL INDICATE YOUR AGREEMENT TO THEM. IF YOU DO NOT AGREE TO ALL OF THE TERMS
AND CONDITIONS IN THIS AGREEMENT, PKI IS NOT WILLING TO LICENSE THE SOFTWARE TO YOU, AND YOU (1) MAY NOT USE
THE SOFTWARE AND (2) MUST CONTACT PKI WITHIN THREE (3) DAYS OF RECEIPT TO ARRANGE FOR THE RETURN OF THE
SOFTWARE. BY INSTALLING AND/OR USING THE SOFTWARE, YOU ALSO REPRESENT AND WARRANT THAT YOU HAVE THE
AUTHORITY TO AGREE TO AND BE BOUND BY THE TERMS OF THE AGREEMENT.
1 Single Use License. The Software is licensed to you and not sold, and you acknowledge that there is no transfer of title or ownership of
the Software to you. Subject to the terms and conditions of this Agreement and your payment of the license fees for the Software, PKI
hereby grants to you a restricted, non-exclusive, non-transferable, non-assignable, non-sublicensable and revocable single-use license to
use, for your internal research purposes only, the executable code version of the Software and the accompanying documentation in hard
copy or electronic format. PKI grants no other licenses hereunder, except for the Research Use Only Vectra Polaris Software set forth
above. Nothing in this Agreement shall be construed as a general internal use license. For the avoidance of doubt, no right to use Vectra
Polaris in Diagnostic Applications is granted under this Agreement. PKI RESERVES ALL RIGHTS NOT EXPRESSLY GRANTED BY THIS
AGREEMENT.
2 Specific Restrictions. You may use the Software only on a single computer. You may make only one (1) copy of the Software, solely for
backup purposes, and such copy must reproduce and include all copyright notices indicating PKI's ownership of the Software. Except as
expressly permitted by applicable law or PKI, you agree that neither you nor a third party acting on your behalf, directly or indirectly, shall:
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covenants contained in this Agreement are reasonable and necessary to protect the legitimate interests of PKI, that PKI would not have
entered into this Agreement in the absence of such covenants, and that your breach or threatened breach of such covenants shall cause
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covenants and for any other relief such court deems appropriate. This right shall be in addition to any other remedy available to PKI at law
or in equity.
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
PerkinElmer Software License Agreement 98
4 Termination. The license granted in Section 1 above is effective until terminated. This Agreement is conditioned upon your continued
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5 Limited Warranty. The sole and exclusive warranty applicable to software and firmware products provided by PKI for use with a
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6 No Other Warranties. EXCEPT FOR THE LIMITED WARRANTY STATED IMMEDIATELY ABOVE AND TO THE MAXIMUM EXTENT
PERMITTED BY APPLICABLE LAW, THE SOFTWARE IS PROVIDED “AS IS” WITHOUT WARRANTY OF ANY KIND, AND PKI
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No modification or waiver of any provision of this Agreement shall be effective unless in writing and signed by the parties. No delay or
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event any provision of this Agreement is held to be unenforceable, the remaining provisions of this Agreement will remain in full force and
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unlawful purpose nor export or re-export the Software except as authorized by law.
r the amount you
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
PerkinElmer Software License Agreement 99
10 TIFF module: This software uses the LibTIFF 4.0.6 library (http://remotesensing.org/libtiff/)
Copyright (c) 1988-1997 Sam Leffler Copyright (c) 1991-1997 Silicon Graphics, Inc.
Permission to use, copy, modify, distribute, and sell this software and its documentation for any purpose is hereby granted without fee,
provided that (i) the above copyright notices and this permission notice appear in all copies of the software and related documentation,
and (ii) the names of Sam Leffler and Silicon Graphics may not be used in any advertising or publicity relating to the software without the
specific, prior written permission of Sam Leffler and Silicon Graphics.
THE SOFTWARE IS PROVIDED “AS-IS” AND WITHOUT WARRANTY OF ANY KIND, EXPRESS, IMPLIED OR OTHERWISE,
INCLUDING WITHOUT LIMITATION, ANY WARRANTY OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE.
IN NO EVENT SHALL SAM LEFFLER OR SILICON GRAPHICS BE LIABLE FOR ANY SPECIAL, INCIDENTAL, INDIRECT OR
CONSEQUENTIAL DAMAGES OF ANY KIND, OR ANY DAMAGES WHATSOEVER RESULTING FROM LOSS OF USE, DATA OR
PROFITS, WHETHER OR NOT ADVISED OF THE POSSIBILITY OF DAMAGE, AND ON ANY THEORY OF LIABILITY, ARISING OUT
OF OR IN CONNECTION WITH THE USE OR PERFORMANCE OF THIS SOFTWARE.
11 This software uses the Vigra 1.6.0 library (http://hci.iwr.uni-heidelberg.de/vigra/)
This software includes machine-executable object code generated by a source language processor from the Vigra libraries covered by
the VIGRA license. These libraries have not been modified by PKI and all rights are reserved by the copyright holder. This software is
provided “as is” without express or implied warranty, and with no claim as to its suitability for any purpose.
The Vigra library is Copyright © 2008, Ulrich Kothe, Heidelberg Collaboratory for Image Processing
The VIGRA License
=================
(identical to the MIT X11 License)
Permission is hereby granted, free of charge, to any person obtaining a copy of this software and associated documentation files (the
“Software”), to deal in the Software without restriction, including without limitation the rights to use, copy, modify, merge, publish,
distribute, sublicense, and/or sell copies of the Software, and to permit persons to whom the Software is furnished to do so, subject to the
following conditions:
The above copyright notice and this permission notice shall be included in all copies or substantial portions of the Software.
THE SOFTWARE IS PROVIDED “AS IS”, WITHOUT WARRANTY OF ANY KIND EXPRESS OR IMPLIED, INCLUDING BUT NOT
LIMITED TO THE WARRANTIES OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE AND NONINFRINGEMENT. IN
NO EVENT SHALL THE AUTHORS OR COPYRIGHT HOLDERS BE LIABLE FOR ANY CLAIM, DAMAGES OR OTHER LIABILITY,
WHETHER IN AN ACTION OF CONTRACT, TORT OR OTHERWISE, ARISING FROM, OUT OF OR IN CONNECTION WITH THE
SOFTWARE OR THE USE OR OTHER DEALINGS IN THE SOFTWARE.
12 This software uses the Boost 1.53.0 library (http://www.boost.org/)
This software includes machine-executable object code generated by a source language processor from the Boost libraries covered by
the Boost license. These libraries have not been modified by PKI and all rights are reserved by the copyright holder. This software is
provided “as is” without express or implied warranty, and with no claim as to its suitability for any purpose.
13 This software uses the Quan 1.0.0 library (https://lists.sourceforge.net/lists/listinfo/quan-matters)
This software includes machine-executable object code generated by a source language processor from the Quan libraries covered by
the Boost license. These libraries have not been modified by PKI and all rights are reserved by the copyright holder. This software is
provided “as is” without express or implied warranty, and with no claim as to its suitability for any purpose.
The Boost license is reproduced below:
Boost Software License - Version 1.0 - August 17th, 2003
Permission is hereby granted, free of charge, to any person or organization obtaining a copy of the software and accompanying
documentation covered by this license (the “Software”) to use, reproduce, display, distribute, execute, and transmit the Software, and to
prepare derivative works of the Software, and to permit third-parties to whom the Software is furnished to do so, all subject to the
following:
The copyright notices in the Software and this entire statement, including the above license grant, this restriction and the fo
disclaimer, must be included in all copies of the Software, in whole or in part, and all derivative works of the Software, unless such copies
or derivative works are solely in the form of machine-executable object code generated by a source language processor.
THE SOFTWARE IS PROVIDED “AS IS”, WITHOUT WARRANTY OF ANY KIND, EXPRESS OR IMPLIED, INCLUDING BUT NOT
LIMITED TO THE WARRANTIES OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE, TITLE AND NONINFRINGEMENT. IN NO EVENT SHALL THE COPYRIGHT HOLDERS OR ANYONE DISTRIBUTING THE SOFTWARE BE LIABLE FOR
ANY DAMAGES OR OTHER LIABILITY, WHETHER IN CONTRACT, TORT OR OTHERWISE, ARISING FROM, OUT OF OR IN
CONNECTION WITH THE SOFTWARE OR THE USE OR OTHER DEALINGS IN THE SOFTWARE.
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14 This software uses the libjpeg-turbo 1.5.0 library (http://libjpeg-turbo.virtualgl.org)
This software makes use of the libjpeg-turbo library. This software is based in part on the work of the Independent JPEG Group. This
library has not been modified by PKI and all rights are reserved by the copyright holder. This software is provided “as is” without express
or implied warranty, and with no claim as to its suitability for any purpose. The library is covered by IJG License. Additionally libjpeg-turbo
incorporates parts of libjpeg which is covered by the BSD license.
The IJG license is reproduced below:
The authors make NO WARRANTY or representation, either express or implied, with respect to this software, its quality, accuracy,
merchantability, or fitness for a particular purpose. This software is provided “AS IS”, and you, its user, assume the entire risk as to its
quality and accuracy.
This software is copyright (C) 1991-1998, Thomas G. Lane. All Rights Reserved except as specified below.
Permission is hereby granted to use, copy, modify, and distribute this software (or portions thereof) for any purpose, without fee, subject
to these conditions:
(1) If any part of the source code for this software is distributed, then this README file must be included, with this copyright and no-
warranty notice unaltered; and any additions, deletions, or changes to the original files must be clearly indicated in accompanying
documentation.
(2) If only executable code is distributed, then the accompanying documentation must state that “this software is based in part on the work
of the Independent JPEG Group”.
(3) Permission for use of this software is granted only if the user accepts full responsibility for any undesirable consequences; the authors
accept NO LIABILITY for damages of any kind.
These conditions apply to any software derived from or based on the IJG code, not just to the unmodified library. If you use our work, you
ought to acknowledge us.
Permission is NOT granted for the use of any IJG author's name or company name in advertising or publicity relating to this software or
products derived from it. This software may be referred to only as “the Independent JPEG Group's software”.
We specifically permit and encourage the use of this software as the basis of commercial products, provided that all warranty or liability
claims are assumed by the product vendor.
ansi2knr.c is included in this distribution by permission of L. Peter Deutsch, sole proprietor of its copyright holder, Aladdin Enterprises of
Menlo Park, CA. ansi2knr.c is NOT covered by the above copyright and conditions, but instead by the usual distribution terms of the Free
Software Foundation; principally, that you must include source code if you redistribute it. (See the file ansi2knr.c for full details.) However,
since ansi2knr.c is not needed as part of any program generated from the IJG code, this does not limit you more than the foregoing
paragraphs do.
The Unix configuration script “configure” was produced with GNU Autoconf. It is copyright by the Free Software Foundation but is freely
distributable. The same holds for its supporting scripts (config.guess, config.sub, ltconfig, ltmain.sh). Another support script, install-sh, is
copyright by M.I.T. but is also freely distributable.
It appears that the arithmetic coding option of the JPEG spec is covered by patents owned by IBM, AT&T, and Mitsubishi. Hence
arithmetic coding cannot legally be used without obtaining one or more licenses. For this reason, support for arithmetic coding has been
removed from the free JPEG software. (Since arithmetic coding provides only a marginal gain over the unpatented Huffman mode, i
unlikely that very many implementations will support it.) So far as we are aware, there are no patent restrictions on the remaining code.
The IJG distribution formerly included code to read and write GIF files. To avoid entanglement with the Unisys LZW patent, GIF reading
support has been removed altogether, and the GIF writer has been simplified to produce “uncompressed GIFs”. This technique does not
use the LZW algorithm; the resulting GIF files are larger than usual, but are readable by all standard GIF decoders.
We are required to state that “The Graphics Interchange Format(c) is the Copyright property of CompuServe Incorporated. GIF(sm) is a
Service Mark property of CompuServe Incorporated.”
The BSD for libjpeg license is reproduced below:
The authors make NO WARRANTY or representation, either express or implied, with respect to this software, its quality, accuracy,
merchantability, or fitness for a particular purpose. This software is provided “AS IS”, and you, its user, assume the entire risk as to its
quality and accuracy.
This software is copyright (C) 1991-2010, Thomas G. Lane, Guido Vollbeding.
All Rights Reserved except as specified below.
Permission is hereby granted to use, copy, modify, and distribute this software (or portions thereof) for any purpose, without fee, subject
to these conditions:
(1) If any part of the source code for this software is distributed, then this README file must be included, with this copyright and no-
warranty notice unaltered; and any additions, deletions, or changes to the original files must be clearly indicated in accompanying
documentation.
t is
P/N CLS147553 Rev. G Vectra Polaris User Manual PerkinElmer, Inc.
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