LEICA TCS SP5 User Manual
Leica TCS SP5
User Manual
Leica Microsystems CMS GmbH
Am Friedensplatz 3
68165 Mannheim
Germany
Telefon: +49 621 7028 0
Fax: +49 621 7028 2980
E-Mail: CLSM.support@leica-microsystems.com
1 Notes about this manual page 1.............
1.1 Copyright page 1..............................................
2 General page 3...........................
2.1 About these operating instructions page 3.........................
3 Proper intended use page 5.................
4 Legal Notes page 7........................
4.1 DISCLOSURE page 7..........................................
4.2 REVISIONS AND CHANGES page 8.............................
4.3 WARRANTY page 8............................................
4.4 Patents page 8................................................
4.5 TRADEMARKS page 10.........................................
Table of Contents
4.6 Software licenses page 10.......................................
5 TCS SP5 specifications page 11..............
5.1 System overview page 11........................................
5.2 Dimensions page 12.............................................
5.2.1 System with inverted stand: page 12...............................
5.2.2 System with upright stand page 13................................
5.3 Installation site requirements page 13..............................
5.4 Permissible ambient conditions page 14............................
5.5 Electrical connections page 14....................................
5.6 Required cooling capacity page 16................................
5.7 Important additional notes page 16................................
5.8 Required safety measures for VIS and UV systems page 17..........
5.9 Required laser safety measures for MP systems page 17............
5.10 Important additional notes page 17................................
6 Safety Notes page 19.......................
6.1 Which standards does this product meet? page 19..................
6.2 Which laser class does the product have? page 21..................
6.3 Laser class for VIS and UV systems page 21.......................
6.4 Laser class for MP systems page 21..............................
6.5 Warnings, Safety Cautions, and Notes page 22.....................
6.6 What should the user of the laser scanning microscope observe? page 23
6.7 Safety Notes for the User page 25................................
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Table of Contents
6.8 Safety Notes for Operation page 26...............................
6.9 Specific Safety Notes page 30....................................
6.9.1 Operational reliability page 30....................................
6.9.2 De-energizing the system page 30................................
6.9.3 Maximum current load of the power outlet strip at the supply unit page 31
6.10 Laser safety devices page 32.....................................
6.10.1 Eye protection for VIS or UV systems page 32......................
6.10.2 Detachable-key switch page 33...................................
6.10.3 Emissions warning indicators page 34.............................
6.10.4 Remote interlock connector on the supply unit page 36...............
6.10.5 Remote interlock connector on the external 405nm advanced laser page 37
6.10.6 Remote interlock connectors on additional external lasers page 37....
6.10.7 Interlock connection on scanner page 39...........................
6.10.8 Function and position of safety switches page 40....................
6.10.9 Transmitted-light lamp housing for inverted stands page 40...........
6.10.10 Transmitted-light lamp housing for upright stands page 42............
6.10.11 Mirror housing on upright stand page 43...........................
6.10.12 Special laser safety equipment page 44............................
6.10.12.1 Safety beam guide and beam collector page 44.....................
6.10.12.2 Eye protection for MP systems page 45............................
6.10.12.3 Shielding in MP systems (UV laser) page 46.......................
6.11 Overview of usable VIS/UV lasers page 47.........................
6.12 Overview of usable MP lasers (IR) page 48.........................
6.13 Safety label on TCS SP5 system page 49..........................
6.13.1 Inverted stand DMI 4000/6000 CS page 49.........................
6.13.2 Upright stand DM 5000/6000 CS page 50..........................
6.13.3 Scan head page 53.............................................
6.13.4 Supply unit page 54.............................................
6.13.5 External 405-nm laser Advance / 405-nm imaging laser page 55......
6.13.6 External 488-nm laser page 56...................................
6.13.7 MP beam coupling unit page 57...................................
6.13.8 Cover (for replacement flange) page 58............................
6.14 Requirements related to the installation/storage location page 59......
6.15 Changing the installation site page 60..............................
6.16 Scanner cooling page 61.........................................
7 Care and cleaning page 63..................
7.1 Cleaning the optical system of the microscope page 63..............
7.2 Cleaning the microscope surface page 64..........................
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8 Startup of the system page 67................
8.1 Setting Up Users page 70........................................
9 Starting the LAS AF page 75................
10 Introduction to LAS AF - Help page 79........
10.1 General page 79................................................
10.1.1 Calling Online Help page 79......................................
10.1.2 Structure of the online help page 80...............................
10.2 Imprint page 81.................................................
10.2.1 In the respective context (context-sensitive) page 82.................
10.2.2 Via the Help menu page 82.......................................
10.2.3 Full-text search with logically connected search terms page 83........
10.3 Key combinations page 84.......................................
Table of Contents
11 Structure of the user interface page 85........
1 1.1 General structure of the user interface page 85.....................
12 What is the Köhler illumination? page 87......
12.1 Setting the Köhler Illumination page 88............................
13 Introduction to confocal work page 91........
13.1 Preparation page 91.............................................
13.1.1 The objective page 92...........................................
13.1.2 Conventional microscopy page 93.................................
13.1.3 Why scan? page 96.............................................
13.1.4 How is an optical section created? page 98.........................
13.2 Acquiring optical sections page 100................................
13.2.1 Data acquisition page 101.........................................
13.2.2 Illumination p age 103.............................................
13.2.3 Beam splitting page 104..........................................
13.2.4 Emission bands page 104.........................................
13.2.5 The pinhole and its effects page 106................................
13.2.6 Image detail and raster settings page 108...........................
13.2.7 Signal and noise page 113........................................
13.2.8 Profile cuts page 116.............................................
13.3 Multiparameter fluorescence page 117..............................
13.3.1 Illumination p age 1 17.............................................
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Table of Contents
13.3.2 Beam splitting page 118..........................................
13.3.3 Emission bands page 119.........................................
13.3.4 Crosstalk page 119...............................................
13.3.5 Sequential capture page 121......................................
13.3.6 Unmixing page 121...............................................
13.4 3D Series page 124..............................................
13.4.1 z-stack page 124.................................................
13.4.2 About section thickness... page 124................................
13.4.3 ...and spacing page 125...........................................
13.4.4 Data volumes page 125...........................................
13.4.5 Depictions page 127..............................................
13.4.5.1 Gallery page 127.................................................
13.4.5.2 Movie page 128..................................................
13.4.5.3 Orthogonal projections page 128...................................
13.4.5.4 Rotated projections page 129......................................
13.5 Time series page 131.............................................
13.5.1 About scan speeds page 131......................................
13.5.2 Points page 132.................................................
13.5.3 Lines page 132..................................................
13.5.4 Planes page 132.................................................
13.5.5 Spaces (time-space) page 133.....................................
13.5.6 FRAP measurements page 133....................................
13.6 Spectral series page 134..........................................
13.6.1 Data acquisition and utilization page 134............................
13.6.2 About spectral resolution page 134.................................
13.7 Combinatorial analysis page 135...................................
5 Switching off the system page 137.............
6 Contact page 139...........................
7 Declaration of conformity page 141............
8 Glossary page 143..........................
18 Safety data sheets page 153...................
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1 Notes about this manual
1.1 Copyright
The contents of this manual are current as of
thetimeofpublicationandaresubjectto
change without notice.
Leica Microsystems CMS GmbH assumes
no warranty with respect to this manual, including but not limited to implicit warranties for
salability and for suitability for a specific purpose. Leica Microsystems CMS GmbH is not
liable for errors or accidental damages or incidental damages in conjunction with equipment, performance or usage of this manual
or the examples contained therein.
Notes about this Manual
© Leica Microsystems CMS GmbH
This document contains information that is
protected by copyright law. All rights reserved. Duplication, adaptation or translation of
this manual requires the prior written approval
of Leica Microsystems CMS GmbH unless
otherwise permitted through copyright arrangements.
The programs for controlling this product are
protected by copyright law. All rights reserved. Reproduction, adaptation or translation
of these programs w ithout prior written approval of Leica Microsystems CMS GmbH is
strictly prohibited.
MicrosoftWindows,WindowsXP,theWindows logo and the Windows CE logo are ei-
User Manual TCS SP5
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Page 1
Notes about this Manual
ther registered trademarks or trademarks of
Microsoft Corporation in the US and/or other
countries. Microsoft products are licensed for
other companies by Microsoft Licensing, Inc.,
a subsidiary company that is wholly owned
by Microsoft Corporation.
All other tradenames and product names in
this document are brands, service brands,
trademarks or registered trademarks of their
respective owners.
Page 2
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2 General
2.1 About these operating instructions
The main focus of these operating instructions is directed to the safety instructions that
must be observed while working with the laser scanning microscope.
In addition, these operating instructions provide a rough overview of the operating principle of laser scanning microscopes. It presents you with the first steps for starting and
startup of the system and provides a description of the Leica Application Suite Advanced
Fluorescence (LAS AF).
General
The Leica TCS SP5 is supplied with the latest version of the licensed Leica Application
Suite Advanced Fluorescence. To maintain
information on the most current level, the
description of software functions was inten tionally omitted from these operating instructions. Instead, you are referred to the online
help of the Leica Application Suite Advanced
Fluorescence in which you can obtain the
most up-to-date explanations and instructions about the corresponding software functions.
Please read the chapter ”Introduction to the
help of the Leica Application Suite Advanced
Fluorescence” in these operating instructions
to familiarize yourself first with setup and
operation. Additional information about specific functions can subs equently be obtained
electronically at your TCS workstation.
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Page 3
General
Page 4
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3 Proper intended use
The system was designed for confocal recording (laser scanning images) of fluorescence-marked living and fixed specimens
as well as for quantitative measurements in
the area of life science.
This system is intended for use in a lab.
Applications of in-vitro diagnostics according
to the medical products law are excluded
from proper intended use.
The manufacturer assumes no liability for
any improper intended use and for use outside the specifications of Leica Microsystems
CMSGmbHaswellasanyrisksresulting
from it. In such cases, the declaration of conformity becomes invalid.
Proper Intended Use
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Page 5
Proper Intended Use
Page 6
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4LegalNotes
Legal Notes
Manufactured in the Federal Republic of
Germany.
© Copyright 2005,
Leica Microsystems CMS GmbH.
All rights reserved.
No part of this publication may be reproduced or transmitted in any form or by any
means, electronic or mechanical, including
photocopying, recording, or storing in a retrieval system, or translating into any language in any form without the express written permission of Leica Microsystems CMS
GmbH. This also includes photocopying, recording or storing on a retrievable system as
well as the translation into another language.
4.1 DISCLOSURE
This document contains Leica Microsystems
CMS GmbH proprietary data and is provided
solely to its customers for their express benefit of safe, efficient operation and maintenance of the product described herein.
Use or disclosure of Leica Microsystems
CMS GmbH proprietary data for the purpose
of manufacture or reproduction of the item
described herein, or any similar item, is prohibited, and delivery of this document shall
not constitute any license or implied authorization to do so.
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Legal Notes
4.2 REVISIONS AND CHANGES
Leica Microsystems CMS GmbH reserves
the right to revise this document and/or to
further develop and improve the products
described in this document at any time without prior notice or any other obligation.
Information and specifications in this manual
are subject to change without notice.
4.3 WARRANTY
Leica Microsystems CMS GmbH provides
this publication ”as is” without warranty of
any kind, either expressed or implied, including but not limited to the implied warranties
of merchantability or fitness for a particular
purpose.
All reasonable precautions have been taken
in the preparation of this document, including
both technical and non-technical proofing.
Leica Microsystems CMS GmbH accepts no
responsibility for any errors or omissions. In
addition, Leica Microsystems CMS GmbH is
not responsible for direct, incidental or consequential damages that could result from
the use of the material described in this document.
4.4 Patents
Page 8
The product is protected by the following US
patents:
5,886,784; 5,903,688; 6,137,627; 6,222,961;
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Legal Notes
6,285,019; 6,311,574; 6,355,919; 6,423,960;
6,433,814; 6,444,971; 6,466,381; 6,510,001;
6,614,526; 6,654,165; 6,657,187; 6,678,443;
6,687,035; 6,738,190; 6,754,003; 6,801,359;
6,831,780; 6,850,358; 6,867,899.
Further patents are pending.
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Page 9
Legal Notes
4.5 TRADEMARKS
4.6 Software licenses
Throughout this manual, trademarked names
may be used. Rather than including a trademark (™) symbol at every occurrence of a
trademarked name, we state that we are
using the names only in an editorial fashion,
and to the benefit of the trademark owner,
with no intention of infringement.
The software described in this document is
furnished under a License Agreement which
is included with the product. This agreement
specifies the permitted and prohibited uses
of the product.
Page 10
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5 TCS SP5 specifications
5.1 System overview
12
Specifications
6
figure1. System components (overview)
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5
1 TCS SP5 Scanner
2 Control panel
3 TCS workstation
4 Supply unit
5 Panel box
6 Stand with scanner
4
3
Page 11
Specifications
5.2 Dimensions
5.2.1 System with inverted stand:
figure2. Dimensions of the TCS SP5 with inverted stand
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5.2.2 System with upright stand
Specifications
figure3. Dimensions of the TCS SP5 with upright stand
5.3 Installation site requirements
Do not expose the system to draft.
Ensure that the system is not installed next
to elevators, air conditioners or ventilation
systems. For this reason, the installation site
should be carefully planned.
Ensure that the e nvironment is as dustfree as possible.
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Specifications
Please also read the notes on protection
against dust in the chapters on maintenance
and cleaning. Installing the system in darkened rooms is also advisable.
For installation, maintenance and transport, the TCS SP5 system requires doors
with inside spans of 1.00 m.
With regard to the load-bearing capacity of
the floor, please note that the system will apply a static load of 200 kg/m@.
5.4 Permissible ambient conditions
Permissible temperature range for operation:
18 ... 25° C
Permissible relative humidity: 20-80% (noncondensing)
Permissible vibration speeds:
Frequency range [5 Hz–30 Hz]: less than 30
μm/s (effective value)
Frequency range [> 30 Hz]: less than 60
μm/s (effective value)
Pollution degree: Class 2
5.5 Electrical connections
Supply voltage (Input 1/Input 2): 100...240 V
AC +/- 10%
Frequency: 50/60 Hz
Page 14
Power consumption: 2 x 1.6kW
Overvoltage category: II
The building installation must feature two se-
parate power connections with the following
fuse protection:
D 100 V - 120 V AC power: 20 A
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Specifications
D 200 V - 240 V AC power: 12-16 A
For the specifications of external lasers such
as UV and MP lasers, please refer to the manufacturer’s documentation.
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Specifications
5.6 Required cooling capacity
The TCS SP5 system has a maximum power
consumption of 3.2
kW (VIS system).
For the specifications of external lasers such
as UV and MP lasers, please refer to the manufacturer’s documentation.
5.7 Important additional notes
The optimal optical performance of the system can be achieved using standard objectives and standard immersions only at 22°
Celsius + 1 Kelvin.
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Specifications
5.8 Required safety measures for VIS and UV systems
Please observe valid country-specific regulations pertaining to laser safety measures for
laser class IIIb. The operator is responsible
for observing the laser safety regulations.
5.9 Required laser safety measures for MP systems
Please observe valid country-specific regulations pertaining to laser safety measures for
laser class 4/IV. The operator is responsible
for observing the laser safety regulations.
5.10 Important additional notes
The optimal optical performance of the systemcanbeachievedusing standard objectives and standard immersions only at 22°
Celsius + 1 °Celsius.
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Specifications
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6 Safety Notes
Safety Notes
The instrument is Class 3B or 4 laser
equipment
(depending on the laser used) in accordance with IEC/EN 60225-1.
This laser equipment may be operated
only by persons who have been trained in
the use of the systems and the potential
dangers of laser radiation.
As it is not possible to anticipate every potential hazard, please take care and apply
common sense to the installation, operation
and maintenance of this product. Observe all
safety precautions relevant to Class IIIb lasers and Class IV MP systems.
Do not deviate from the operating and maintenance instructions provided herein.
The failure to observe these instructions
shall be exclusively at the user’s own risk
and may void the warranty.
6.1 Which standards does this product meet?
This device was tested and meets the requirements of the following standards:
D IEC/EN 61010-1
”Safety requirements for electrical equipment for measurement, control and laboratory use”
D IEC/EN 60825-1
”Safety of laser products, Part
1: Equipment classification, requirements and user’s guide”
D IEC/EN 61326
”Electrical equipment for measurement,
control and laboratory use - EMC requirements” (Class A).
This is a Class A device. Operating this
device on a public low-voltage grid may
result in radio interference. The operator
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Safety Notes
must take suitable measures should this
occur.
D IEC/EN 61000-3-2 (EMC)
”Electromagnetic Compatibility” Part
3-2: Limits - Limits for harmonic currents
D IEC/EN 61000-3-3 (EMC)
”Electromagnetic Compatibility” Part
3-3: Limits - Limits - Section
3: Limits for voltage fluctuations and flikker in low-voltage networks.
D For use in the USA:
CDRH 21 CFR 1040.10:
Laser Products U.S. Food and Drug
Administration (FDA) (”Complies with
FDA performance standards for laser
products except for deviations pursuant
to laser notice No. 50, dated 26 July,
2001.”)
For the scope of the CDRH/FDA
(USA), the designation Laser Class
3B in the text must be replaced by IIIb
andClass4byIV.
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Safety Notes
6.2 Which laser class does the product have?
Laser
type
VIS 400 - 700 nm,
UV 350 - 700 nm,
MP 350 - 1050 nm,
Wavelength range Configuration Laser class
Combination of lasers
(visible laser
radiation)
(visible and invisible la-
ser radiation)
(visible and invisible la-
ser radiation)
from Chapter 6.11
(without lasers having wave-
lengths of 350 - 400 nm)
Combination of lasers
from Chapter 6.11
(VIS and UV lasers)
Combination of lasers from
Chapter 6.11 (VIS/UV lasers)
and Chapter 6.12 (IR lasers)
6.3 Laser class for VIS and UV systems
According to IEC/EN 60825-1, this laser
scanning microscope is a laser device of
Class IIIb.
3B / IIIb
3B / IIIb
4/IV
Avoidexposingeyesorskintodirectradiation.
6.4 Laser class for MP systems
According to IEC/EN 60825-1, this laser
scanning microscope is a laser device of
Class IV.
Avoidexposingeyesorskintodirectand
indirect radiation.
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Safety Notes
6.5 Warnings, Safety Cautions, and Notes
Notes either contain additional information
on a specific topic or special instructions on
the handling of the product.
A safety note points out an operation, a process,a condition oran instruction that must be
observed strictly to prevent severe damage to
the system or loss of data.
A laser warning points out an operation, a
process, a condition or an instruction that
must be observed strictly to prevent serious
eye injuries to the persons using the system.
A high-voltage warning points out an operation, a process, a condition or an instruction that must be observed strictly to prevent
possible injury or death of the persons using
the system.
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Safety Notes
6.6 What should the user of the laser scanning microscope observe?
The user of this product is responsible
for proper and safe operation and safe
maintenance of the system as well as for
following all applicable safety regulations. The user is
fully liable for all consequences resulting
from the use of the system for any other purposes than those listed in the Operating Instructions or the online help.
The user is obligated to perform and monitor suitable safety measures
according to IEC / EN 60825-1 and
the corresponding national regulations.
Users must have received instructions
concerning the risk potential associated with
the operation of laser devices.
To assure classification as a
3B/IIIb or 4/IV laser product according to
IEC/EN60825-1 and electrical safety compliance, all safety devices, interlocks, and
safety systems of the laser device must
be in operational condition.
Deactivating or damaging these safety devices or any intervention in any of these safety
devices may lead to serious eye injuries,
physical injuries or property damages. In
these cases, Leica Microsystems CMS
GmbH does not assume any liability.
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According to IEC/EN 60825-1:
”Safety of laser products, Part
1: Equipment classification, requirements
and user’s guide,” the user is required to
Page 23
Safety Notes
designate a Laser Safety Officer or a Laser Protection Advisor.
Repairs and servicing may only be performed by authorized Leica Microsystems
CMS GmbH service personnel.
The user is fully liable for all
consequences resulting from the use of the
system if it is opened, improperly serviced or
repaired by other persons than authorized
Leica customer service representatives.
If repairs or service measures are performed that require opening parts of the
housing, only trained Leica service technicians may occupy the room in which
the laser scanning microscope is located.
Leica Microsystems CMS GmbH will not be liable for damages resulting from nonobservance of the
above information. The above information
does not, in any way, implicitly or explicitly,
modify the warranty and liability clauses contained in the general terms and conditions of
Leica Microsystems CMS GmbH.
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