Thermo Scientific PikoReal
Real-Time PCR System
User Manual
Rev. 2.0
Thermo Scientific PikoReal
Real-Time PCR System
User Manual
Rev.2.0, Cat. no. N11471
Copyright
Copyright © 2011 Thermo Fisher Scientific Corporation. All rights reserved. Reproduction of the
accompanying user documentation in whole or in part is prohibited.
Licensing information
Not available in Canada, Brazil, UK, Germany, Austria, Switzerland, Italy, Spain, France, Belgium, the
Netherlands, Liechtenstein, Denmark and Sweden.
Notice: Purchase of this instrument conveys a limited, non-transferable immunity from suit for the purchaser's
own internal research and development and applied fields other than human in vitro diagnostics under one or
more of Canadian Patent 1,339,653, U.S. Patent 5,475,610 (claims 160-163 only) or corresponding claims in
their unexpired non-U.S. counterparts owned by Applied Biosystems. No right is conveyed expressly, by
implication or by estoppel under any other patent claim, including under U.S. Patent No. 6,814,934 or any
global equivalent thereof, which claims thermal cycler apparatus and systems capable of real-time detection.
Further information on purchasing licenses may be obtained by contacting the Director of Licensing, Applied
Biosystems, 850 Lincoln Centre Drive, Foster City, California, 94404, USA.
Trademarks
“ALPS”, “DyNAmo”, “Finnpipette”, “Matrix”, “Maxima”, “Multidrop”, “Piko”, “PikoReal”, “Solaris”,
“Versette”, “Verso” and “UTW” are trademarks or registered trademarks of Thermo Fisher Scientific Inc. and
its subsidiaries.
All other trademarks and registered trademarks are the property of their respective holders.
Disclaimer
Thermo Fisher Scientific reserves the right to change its products and services at any time to incorporate
technological developments. This manual is subject to change without prior notice as part of a continuous
product development. Although this manual has been prepared with every precaution to ensure accuracy,
Thermo Fisher Scientific assumes no liability for any errors or omissions, nor for any damages resulting from
the application or use of this information. This manual supersedes all previous editions.
Remarks on screenshots
Screenshots may be slightly different on your system depending on the firmware version.
No liability for consequential damages
Thermo Fisher Scientific shall not be liable for any damages whatsoever arising out of the use or inability to
use this product.
Power failure
The system requires uninterrupted power supply in order to operate correctly. Thermo Fisher Scientific has no
responsibility whatsoever for system malfunctions arising from power failures.
About This User Manual
Intended users
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 3
About This User Manual
The PikoReal Real-Time PCR System, as a standalone instrument or
with Thermo Scientific PikoReal Software, can be used in research
laboratories by professional personnel.
This user manual is for the following instruments: 24-well PikoReal
Real-Time PCR System (Cat. no. TCR0024) and 96-well PikoReal
Real-Time PCR System (Cat. no. TCR0096). It aims to give you the
information you need for:
● Reviewing safety precautions
● Installing the PikoReal Real-Time PCR System
● Operating the instrument
● Adjusting the settings and servicing the instrument
● Troubleshooting the instrument
This user manual also describes all the features and specifications of the
PikoReal Real-Time PCR System, and provides ordering information
for instruments, spare parts, reagents and consumables.
Read the manual in its entirety before operating the instrument.
Keep the user manual for future reference. The user manual is an
important part of the instrument and should be readily available.
For PikoReal Software related issues, refer to the Thermo Scientific
PikoReal Software User Manual (Cat. no. N12076).
For the latest information on products and services, visit our website:
http://www.thermoscientific.com/pikoreal
In our efforts to provide useful and appropriate documentation, we
would appreciate any comments you may have on this user manual to
your local Thermo Fisher Scientific representative.
Intended users
How to use this
user manual
For more
information
About This User Manual
Safety symbols and markings
4 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
These symbols are intended to draw your attention to particularly
important information and alert you to the presence of hazards.
The following symbols and markings appear on the type label and the
instrument itself.
Power on ▲
Power off ▲
Serial number ▲
Catalog number ▲
Date of manufacture ▲
Consult instructions for use ▲
WEEE symbol This product is required to comply with the European
Union’s Waste Electrical & Electronic Equipment (WEEE) Directive
2002/96/EC. ▲
Two labels with the following texts (Figure 2–3):
ATTENTION!
This is shipping position only! The other hole is operating position,
refer to the manual. ▲
ATTENTION!
Always insert the plug into the Piko first, and then insert the power
cord into the wall outlet. ▲
Safety symbols
and markings
Safety symbols and
markings used on the
PikoReal Real-Time PCR
System
About This User Manual
Safety instructions
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 5
The following symbols and markings appear in this user manual.
Warning Risk of electric shock. ▲
Warning Biohazard risk. ▲
Warning Hot surface, risk of burns. ▲
Warning Risk of injury to the user(s). ▲
Caution Risk of damage to the instrument, other equipment or loss of
performance or function in a specific application. ▲
Note Marks a hint, important information that is useful in the
optimum operation of the system, or an item of interest. ▲
Read the User Manual Operating the PikoReal Real-Time PCR
System without first reading the entire User Manual may constitute a
risk to your health. Only a person capable of handling electrical
equipment should use the PikoReal Real-Time PCR System. NOT
FOR CHILDREN. ▲
Do Not Attempt to Repair the System Do not remove the cover of the
system. Do not try to repair or replace broken components – you put
yourself at risk for electrical shock. Removing the cover or
replacing/removing components will void the warranty. Contact
customer service or your local distributor if your system is not
functioning properly. ▲
Do Not Touch the Sample Block Certain components, including the
sample block and heated lid, will become excessively hot. Touching
these components may cause burns. ▲
Do Not Tamper with Electronics Coming into contact with the
electronics, even when the system is off or unplugged, may cause an
electrical shock or harm. ▲
Do Not Use Flammable, Hazardous or Radioactive Liquids with the
PikoReal Real-Time PCR System. ▲
W
arning and other
markings used in
the documentation
Safety
instructions
About This User Manual
Safety instructions
6 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
Contents
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 7
Contents
Intended users................................................................................... 3
How to use this user manual ............................................................. 3
For more information .......................................................................3
Safety symbols and markings.............................................................4
Safety symbols and markings used on the PikoReal Real-Time PCR
System............................................................................................ 4
Warning and other markings used in the documentation ............... 5
Safety instructions ............................................................................. 5
Introduction to the PikoReal
®
Real-Time PCR System ........................ 11
Introduction to qPCR..................................................................... 13
Intended use.................................................................................... 13
General Description .................................................................................... 15
Instrument views.............................................................................15
PikoReal user interface .................................................................... 17
Status indicator ............................................................................ 17
Power supply................................................................................... 17
PikoReal Software ........................................................................... 18
Block formats.................................................................................. 18
Consumables................................................................................... 18
UTW technology ......................................................................... 19
Reagents..........................................................................................20
Dye calibration................................................................................ 21
Dispensers....................................................................................... 21
Heat sealers ..................................................................................... 21
Installation .................................................................................................... 23
What to do upon delivery ...............................................................23
Unpacking the instrument ...........................................................23
Checking delivery for completeness and damage ..........................23
Environmental requirements ........................................................ 24
Air flow..................................................................................... 25
Installation steps.............................................................................. 25
Connecting the power supply cable.............................................. 25
Connecting to the network ..........................................................26
Connecting to the data port ......................................................... 26
Warnings and cautions....................................................................26
Electrical ...................................................................................... 26
Operational check ...........................................................................27
Installing PikoReal Software............................................................ 27
Chapter 1
Chapter 2
Chapter 3
Contents
8 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
Routine Operation ........................................................................................29
Switching on ................................................................................... 29
Status indicator ............................................................................... 29
Main menu ..................................................................................... 29
Running a protocol ......................................................................... 30
Monitoring a run ............................................................................32
Status screen................................................................................. 32
Time screen.................................................................................. 33
Functions in the screens ............................................................... 33
Tools menu.....................................................................................34
System info ..................................................................................34
Diagnostics ..................................................................................35
Lid ............................................................................................ 35
Sensors ...................................................................................... 36
Display...................................................................................... 36
Clock ........................................................................................37
Network....................................................................................... 37
Service.......................................................................................... 38
Logfiles......................................................................................... 38
Park instrument ...........................................................................38
Run data to USB..........................................................................39
Shutting down ................................................................................40
Maintenance .................................................................................................41
Regular and preventive maintenance ............................................... 41
Maintenance checklist .................................................................. 41
Instrument care ............................................................................ 41
Cleaning of the instrument .............................................................42
Cleaning the block and wells........................................................ 43
Cleaning the heat sink..................................................................43
Disposal of materials ....................................................................... 44
Decontamination procedure............................................................ 44
Packing for service........................................................................... 45
Maintaining a system log................................................................. 46
Disposal of the instrument .............................................................. 46
Technical Specifications ...........................................................................47
Instrument specifications................................................................. 47
Safety specifications......................................................................... 48
In conformity with the requirements............................................48
Troubleshooting Guide................................................................................ 51
Handling abnormal situations ......................................................... 51
Power failure ...................................................................................51
Warning and error codes................................................................. 51
Chapter 4
Chapter 5
Chapter 6
Chapter 7
Contents
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 9
Ordering Information................................................................................... 55
PikoReal.......................................................................................... 55
List of spare parts and accessories ....................................................55
List of consumables ......................................................................... 56
List of reagents ................................................................................ 57
List of heat sealers ...........................................................................57
System Log .................................................................................................... 59
Certificate of Decontamination................................................................. 61
Chapter 8
Appendix A
Appendix B
Contents
10 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
Introduction to the PikoReal( Real-Time PCR System
Safety instructions
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 11
Chapter 1
Introduction to the PikoReal®
Real-Time PCR System
The PikoReal Real-Time PCR System (Figure 1-1) is a gene
quantification and genotyping platform in 24- and 96-well block
formats. Using PikoReal Software, the system is applicable to absolute
quantification, relative quantification, melt curve analysis and allelic
discrimination. The PikoReal Real-Time PCR System is designed to a
minimal footprint, making the system ideal for personal benchtop use
and field applications, for example.
Figure 1-1. PikoReal 96 Real-Time PCR System
The combination of several innovative technologies applied in design,
including optics, unique heat sink design, block design, plate size and
the usage of special plasticware, create many advantages over
conventional qPCR systems. The PikoReal Real-Time PCR System is
fast, fulfills high-level requirements of temperature uniformity over the
wells and creates significant savings in reagent, plastic material and
energy consumption.
The PikoReal Real-Time PCR System uses ultra-thin wall (UTW®)
microwell plates that are 1/4 of the size of conventional microwell
plates. The plates adhere to industry standards regarding well spacing
and sample capacity and are compatible with multichannel pipettes,
Introduction to the PikoReal( Real-Time PCR System
Safety instructions
12 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
reagent dispensers and automated liquid handling systems. The
PikoReal 24 also utilizes standard low profile 8-well strips and 0.2 ml
tubes.
The optical system comprises a charge-coupled device (CCD) camera,
optical filters and an individual light-emitting diode (LED) for each of
the five channels. The channel selection is built on a motorized linear
shuttle that moves to five channel positions.
The PikoReal is controlled by PikoReal Software via an Ethernet
connection (with up to 10 connections) or alternatively as a standalone
instrument with USB memory stick loaded protocols.
The PikoReal Real-Time PCR System has the following features:
● Compact design – The PikoReal Real-Time PCR System has a
compact design giving a minimal footprint for personal benchtop
use and field applications.
● Unique thermal system – Three Peltier elements and a patented
heat sink design provide a fast and even heat distribution and
removal.
● 5 LED optical system – Separate LEDs for each channel offer a
wide excitation range (475–640 nm) and multiplexing for four
dyes.
● Piko
®
format – The PikoReal Real-Time PCR System sample
blocks (24-well and 96-well) will only accept microplates with a
footprint with the dimensions of a microscope slide – the Piko
format. The 24-well sample block is also compatible with all
standard low-profile 8-tube strips and single tubes.
● Automatic lid – The heated lid is internally motor-controlled and
the lid will automatically adjust to the optimal temperature and
sealing pressure.
● External power supply – The PikoReal Real-Time PCR System
uses an external 200 watt power supply (an approach commonly
used with lap-top computers).
Note Conventional format full-sized microplates are not compatible
with the PikoReal blocks.
▲
The PikoReal Real-Time PCR System is available in two different
configurations:
● PikoReal 24
• Cat. no. TCR0024
• for use with 24-well Piko PCR plates, standard 8-well strips
and 0.2 ml tubes
● PikoReal 96
• Cat. no. TCR0096
Introduction to the PikoReal( Real-Time PCR System
Introduction to qPCR
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 13
• for use with 96-well Piko PCR plates.
The following components form the complete technology platform for
performing real-time PCR based DNA analyses:
● PikoReal Real-Time PCR System
● PikoReal Software (see “PikoReal Software” on page 18)
● UTW® consumables (see “Consumables” on page 18)
● qPCR reagents (see “Reagents” on page 20).
Quantitative real-time PCR (qPCR) is a method for analyzing DNA by
detecting the amount of PCR product after each PCR cycle by the
incorporation of fluorescent dyes into the reaction. The amplification
and detection of a specific DNA sequence is integrated into a
simultaneous process within a single instrument. The product
accumulation is detected and measured throughout the reaction – in
real-time. The detection is based on monitoring fluorescent molecules
that are incorporated into the amplified products as a dsDNA binding
dye or within labeled target specific primers or probes.
The quantitation is based on correlation between the measured
fluorescence intensity and the amount of DNA formed. Collecting
quantitative data of a known dilution series of the target DNA can be
used to give insight into the PCR reaction kinetics and reaction
efficiency. The gene quantification analysis method can be absolute
(copy number, concentration) or relative (to a known reference gene
and calibrator sample). Examples of qualitative analysis include gene
identification and genotyping, e.g. studying DNA sequence variations,
such as single nucleotide polymorphism.
The basic difference from end-point PCR technique is that in addition
to the qualitative result – detecting the presence of a PCR product, a
quantitative result – the initial amount of the amplified product – can
be determined. qPCR offers many advantages over traditional endpoint PCR methods: high sensitivity over a broad dynamic range,
reproducibility, low contamination risk and the exclusion of post-PCR
manipulations for analysis. It also facilitates high-throughput and
multiplexing applications.
The PikoReal Real-Time PCR System is a thermal cycler with a
fluorescence measuring system available in 24-well and 96-well formats.
It is intended for qPCR technologies and related fluorogenic
chemistries. The PikoReal Real-Time PCR System is intended for
research purposes only.
Use for any other than the above-mentioned purposes is excluded.
Introduction to
qPCR
Intended use
Introduction to the PikoReal( Real-Time PCR System
Intended use
14 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
Caution Operate the instrument only with software and hardware that
is specifically designed or selected for it. Thermo Fisher Scientific
assumes no liability for the use of third-party software applications. ▲
General Description
Instrument views
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 15
Chapter 2
General Description
This chapter describes the main parts of the PikoReal instrument.
The front view of the PikoReal instrument is shown in Figure 2–2.
Figure 2–2. PikoReal front view
Instrument
views
Feet
Air intake
Drawer
User interface
Data port
Air exhaust
General Description
Instrument views
16 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
The rear view of the PikoReal instrument is shown in Figure 2–3.
Figure 2–3. PikoReal rear view
Caution Do not block the air intake or the air exhaust vents
(Figure 2–2 and Figure 2–3). Be careful not to place, for example, a
piece of paper under the air intake.
▲
Warning Do not place anything in front of the drawer. Items may be
knocked over when opening the drawer.
▲
Power
connector
Ethernet port (RJ45)
Air exhaust
Type label
General Description
PikoReal user interface
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 17
The PikoReal user interface (UI) and its functions are shown in
Figure 2–4.
Figure 2–4. PikoReal user interface
On the top left corner of the instrument user interface is a status
indicator light that has the following states (Table 2–1):
Table 2–1. Status indicator
Indicator Status
Constant blue Instrument is on but not in use.
Blinking blue Instrument is on and in use.
Constant red System error. For more information, see Chapter 7:
“Troubleshooting Guide”.
The PikoReal Real-Time PCR System is equipped with an external
200 watt power supply.
A green light on the power supply indicates normal operation.
The power supply will get warm during normal operation.
Note If the internal circuit breaker trips, the green light of the power
supply will go off. The power supply must be unplugged from the
power outlet for 5 minutes to reset it.
▲
PikoReal user
interface
Status indicator
Power supply
Open/Close
drawer button
Power button
Backlit LCD display
Arrow keys for
moving on
the display
Cancel key
Enter key
Upper light
Lower light
Status indicator: Blue
Red
General Description
PikoReal Software
18 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
Note Do not cover the power supply with anything as this will impede
the ability of the power supply to efficiently dissipate heat. ▲
The PikoReal is operated with the PikoReal Software application. Up to
ten PikoReals can be simultaneously operated from a single computer.
For more information, refer to the PikoReal Software User Manual
(Cat. no. N12076).
The PikoReal Real-Time PCR instrument is available in two block
formats (Figure 2–5 and Figure 2–6).
Figure 2–5. 24-well, 50 µl max. volume for PCR
Figure 2–6. 96-well, 20 µl max. volume for PCR
PikoReal 24-well and 96-well formats utilize Piko PCR plates, which
are 1/4th the size of standard PCR plates. See Figure 2–7.
The plates maintain the industry standard for well volumes and well
spacing, ensuring the compatibility with standard multi-channel
pipettes (both 8- and 16-channel) and liquid handling robots.
Four 24-well plates are equivalent to a standard ANSI 96-well plate and
four 96-well Piko PCR plates are equivalent to a standard ANSI 384well plate.
PikoReal
Software
Block formats
Consumables
General Description
Consumables
Thermo Fisher Scientific Thermo Scientific PikoReal Real-Time PCR System User Manual 19
Plate frames are available to assemble Piko PCR plates into standard
ANSI format plate size (see Chapter 8: “Ordering Information”).
The 24-well Piko PCR plates are sealed with strip caps or with a heat
sealing film, whereas 96-well Piko PCR plates are sealed with a heat
sealing film.
The 24-well sample block is also compatible with low profile 8-tube
strips (sealed with attached* or separate cap strips) and single tubes with
flat optical caps.
* Not available in US.
Note If you are using single tubes, you must use at least eight tubes that
are placed symmetrically with at least one tube in each corner of the
block.
You can use empty tubes to fill the above requirement when analyzing
fewer samples. ▲
Figure 2–7. Piko PCR plates
Thermo Fisher Scientific Piko PCR plates, strips and tubes utilize
UTW (ultra-thin wall) technology and are recommended to be used
with PikoReal.
UTW reaction vessels were used in the optimization of the temperature
algorithms of the instrument as well as in the validation of its
performance.
Note For the highest signal-to-background ratio and elimination of
possible well-to-well cross-talk, white plates, strips and tubes are
recommended. ▲
UTW technology
General Description
Reagents
20 Thermo Scientific PikoReal Real-Time PCR System User Manual Thermo Fisher Scientific
The UTW consumables are half the thickness (< 0.15mm) of
conventional PCR consumables and thus improve the thermal transfer
between block and sample (see Figure 2–8 and Figure 2–9).
The uniform tube design allows repeatable and consistent results from
well to well.
Figure 2–8. The tube wall thickness of UTW consumables (marked as blue)
is half that of standard consumables (marked as red).
Figure 2–9. The lower portion of the UTW reaction vessels is ultra-thin.
For more information on the consumables, refer to Chapter 8:
“Ordering Information” and http://www.thermoscientific.com/pikoreal.
Thermo Scientific offers a comprehensive selection of qPCR reagents to
be used with the PikoReal Real-Time PCR System. These include, for
example, universal reagents for SYBR Green and probe chemistries,
SNP Genotyping as well as Solaris pre-designed qPCR assays for geneexpression analysis.
Reagents
UTW
area
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