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Zeiss DirectFRAP Operating Manual

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  • Page 2 Persons in contravention of this copyright shall be liable to pay compensation for damages. All rights reserved in the event of a patent grant or registration of a utility patent. Published by: Carl Zeiss Microscopy GmbH Carl-Zeiss-Promenade 10 07745 Jena, Germany microscopy@zeiss.com www.zeiss.com/microscopy...

  • Page 3: Table Of Contents

    System Overview - DirectFRAP Imaging System ..............29 Diaphragm Size and Objective Magnification ..............30 Optical Fibers for the Application of the DirectFRAP Laser Manipulation Slider ....31 Suitability of Lasers for the Application of the DirectFRAP Imaging System ......31 Laser Safety Concept ......................
  • Page 4 Carl Zeiss Contents DirectFRAP Switching on ........................45 During Operation ......................45 Switching off ........................45 Adjusting the Laser Beam with the Laser Manipulation Slider ..........46 Operation via ZEN Software (Blue Edition) ................47 Experiment Procedure ....................... 47 CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE .......... 49 Instrument Care ........................

  • Page 5: Notes On Instrument Safety

    98/79/EC (In- Vitro Diagnostics Directive) and carries the marking. The DirectFRAP imaging system has to be disposed of in compliance with the WEEE Directive 2012/19/EU. The warning and information symbols explained below will help you recognize at a glance the type and...
  • Page 6 Notes on Instrument Safety Carl Zeiss General Notes DirectFRAP In addition to this manual of the DirectFRAP imaging system, please observe the detailed information given in the following operating manuals: Operating manual for the Axio Observer microscope (B 46-0111) −...

  • Page 7: Safety Instructions

    Users should also ensure that they observe all relevant legal requirements. If operated correctly, the DirectFRAP imaging system does not create any risk of radiation exposure for operating personnel. Nevertheless, you should observe the following warnings: Inform the laser safety officer before you begin with procedures for initial start-up of the DirectFRAP imaging system.

  • Page 8: Installation Notes

    Installation Notes The DirectFRAP imaging system must be installed and commissioned by technicians trained in laser safety and authorized by Carl Zeiss. The client is liable to pay the costs of services rendered by Carl Zeiss Microscopy GmbH. To install the DirectFRAP imaging system, ensure a minimum distance of 0.5 m between the wall and the rear of the system.
  • Page 9 Samples should also be disposed of in compliance with the relevant legal requirements and internal operating procedures. The DirectFRAP imaging system must be connected to the power supply using several multiple sockets connected separately to installation sockets. Connection via a single multiple socket plugged into a single installation socket would lead to inadmissibly high contact current levels.

  • Page 10: Notes On Computer And Software

    Never disconnect the power cable while the device is in use. Use the power switch to turn the devices off. To ensure proper function of the DirectFRAP imaging system, it should be subjected to an annual safety inspection. The safety checks must be carried out by an authorized Carl Zeiss service technician.

  • Page 11: Service Interfaces

    Service interfaces between Laser Bench Tower and laser manipulation slider CAUTION The fiber mounts (Fig. 1-1/1 and 2) may be detached by Carl Zeiss Service staff only. If one fiber end is not firmly connected to the fiber mount, laser class 3B radiation will emerge!
  • Page 12 Safety Instructions DirectFRAP Fig. 1-2 Service interfaces between Laser Bench Rack 3F and laser manipulation slider (DirectFRAP), Laser TIRF slider (optional) and Cell Observer SD (optional) CAUTION The fiber mountings in the Laser Bench Rack (Fig. 1-2/1), on the laser manipulation slider (Fig.
  • Page 13 CAUTION The screws (Fig. 1-3/1) on the retaining ring (Fig. 1-3/2) for the camera adapter should only be unscrewed by Carl Zeiss service technicians! Non-observance involves the risk of eye damage! If no camera is mounted on the sideport, it should be sealed by the protective cover (Fig.
  • Page 14 Notes on Instrument Safety Carl Zeiss Safety Instructions DirectFRAP Fig. 1-4 Camera adapter - camera / protective cover customer interface The camera adapter (Fig. 1-4/1) is equipped with an interlock connection (two reflected light barriers) which automatically closes the laser shutter as soon as the camera (Fig. 1-4/2) or the protective cover (Fig.
  • Page 15 CAUTION The screws (Fig. 1-5/1) on the sensor ring (Fig. 1-5/2) and the screw (Fig. 1-5/6) on the contact ring (Fig. 1-5/4) may only be unscrewed by Carl Zeiss service technicians! Only screw (Fig. 1-5/7) may be unscrewed. Non-observance involves the risk of eye damage!
  • Page 16 Transmitted light carrier - HAL lamp service interface CAUTION The clamping screw (Fig. 1-6/1) may be detached by Carl Zeiss Service staff only. If the lamp housing is removed, laser class 3B radiation will emerge! Non-observance involves the risk of eye damage!

  • Page 17: Warning And Information Labels On The Instrument

    Observe the warning and information labels affixed to the DirectFRAP imaging system and Axio Observer microscope. Check that all these labels are attached. If one of the labels is missing, contact Carl Zeiss Microscopy GmbH for a free replacement. You will receive replacements free of charge.
  • Page 18 Notes on Instrument Safety Carl Zeiss Safety Instructions DirectFRAP Fig. 1-8 Warning and information labels on the laser manipulation slider Fig. 1-9 Warning and information labels on the laser bench rack 423635-9001-701 05/2014...
  • Page 19 Notes on Instrument Safety DirectFRAP Safety Instructions Carl Zeiss Fig. 1-10 Warning and information labels on the laser bench tower 05/2014 423635-9001-701...

  • Page 20: Notes On Warranty

    Such actions will render any warranty claims invalid. The customer is not allowed to carry out any maintenance or repair work to the DirectFRAP imaging system apart from that specified in this operating manual. Repairs are only permitted to the Carl Zeiss Service or persons especially authorized by it.

  • Page 21: Instrument Description

    Intended Use Product name: DirectFRAP imaging system The DirectFRAP imaging system is an extension unit for Axio Observer and is used for microscopic analyses of dynamic processes within living cells by short-term local laser radiation with wavelengths from 405 nm to 640 nm.

  • Page 22: Functional Principle

    INSTRUMENT DESCRIPTION Carl Zeiss Functional Principle DirectFRAP Functional Principle The laser manipulation slider makes it possible to couple laser light simultaneously with the conventional fluorescence illumination from the HXP 120 V light source into the reflected-light beam path of Axio Observer.

  • Page 23: Instrument Description And Main Features

    DirectFRAP Instrument Description and Main Features Carl Zeiss Instrument Description and Main Features The following components are necessary for operating the DirectFRAP imaging system in combination with Axio Observer: Axio Observer.Z1 microscope (431007-0000-000, 431007-9901-000 or 431007-9902-000) with the − basic laser safety kit for the Axio Observer.Z1 stand Laser safety device for microscope and peripheral units −...

  • Page 24: Laser Manipulation Slider

    Laser Manipulation Slider The laser manipulation slider is inserted into the reflected-light illumination system of the microscope. The optical design of the DirectFRAP imaging system allows the simultaneous use of the HXP 120 V illuminator and the laser manipulation unit.

  • Page 25: Laser Bench Tower Or Laser Bench Rack

    INSTRUMENT DESCRIPTION DirectFRAP Instrument Description and Main Features Carl Zeiss 2.3.2 Laser Bench Tower or Laser Bench Rack The laser units for the individual laser lines and the corresponding electronics are housed either in the Laser Bench Tower or in the Laser Bench Rack depending on the equipment version selected.

  • Page 26: Signal Distribution Box

    INSTRUMENT DESCRIPTION Carl Zeiss Instrument Description and Main Features DirectFRAP 2.3.3 Signal Distribution Box The system components are connected to the signal distribution box (see operating manual SVB 1). Fig. 2-5 Signal distribution box 423635-9001-701 05/2014...

  • Page 27: Basic Laser Safety Kit For Axio Observer.z1 Stand

    INSTRUMENT DESCRIPTION DirectFRAP Instrument Description and Main Features Carl Zeiss 2.3.4 Basic Laser Safety Kit for Axio Observer.Z1 Stand The basic laser safety kit for the Axio Observer.Z1 stand is composed of an eyepiece shutter (factory- installed or to be installed by Service), laser safety board for the stand, distribution box, and labels.

  • Page 28: Remote Control Switch For Laser Operation

    The laser control button (Fig. 2-7/1) serves to switch laser operation on and off. The Safe (Fig. 2-7/2) and Active (Fig. 2-7/3) LEDs indicate the current laser status of the DirectFRAP laser manipulation slider. Lighting Active and Safe LEDs signalize that laser operation is enabled (all interlocks are safe).

  • Page 29: System Overview - Directfrap Imaging System

    INSTRUMENT DESCRIPTION DirectFRAP System Overview - DirectFRAP Imaging System Carl Zeiss System Overview - DirectFRAP Imaging System Fig. 2-8 System overview 05/2014 423635-9001-701...

  • Page 30: Diaphragm Size And Objective Magnification

    INSTRUMENT DESCRIPTION Carl Zeiss Diaphragm Size and Objective Magnification DirectFRAP Diaphragm Size and Objective Magnification Objective Magnification and Diaphragm Size (µm)* Diaphragm Edge DirectFRAP without TIRF Slider DirectFRAP with TIRF Slider Clockwise Position Type** 100x 100x Operation 11.2 15.1 12.5 19.0...

  • Page 31: Optical Fibers For The Application Of The Directfrap Laser Manipulation Slider

    INSTRUMENT DESCRIPTION DirectFRAP Optical Fibers for the Application of the DirectFRAP Laser Manipulation Slider Carl Zeiss The following objectives are recommended for laser manipulation: Designation Order number 421190-9900-000 Objective α Plan-Fluar 100x/1.45 Oil M27 420792-9800-000 Objective α Plan-Apochromat 100x/1.46 Oil DIC M27 420782-9900-000 Objective Plan-Apochromat 63x/1.40 Oil DIC...

  • Page 32: Laser Safety Concept

    DirectFRAP Laser Safety Concept The DirectFRAP imaging system is equipped with interlocks on the safety-relevant laser components. The interlocks close the laser shutter immediately if one of these components is no longer in a safe condition or is no longer in the correct work position: −...

  • Page 33: Technical Data

    INSTRUMENT DESCRIPTION DirectFRAP Technical Data Carl Zeiss Technical Data Dimensions (width x depth x height) Axio Observer stand ............... approx. 295 mm x 805 mm x 707 mm Laser manipulation slider ................285 mm x 88 mm x 165 mm Signal distribution box ..............
  • Page 34 Carl Zeiss Technical Data DirectFRAP Operating data DirectFRAP overall system: Radio interference suppression ............in compliance with EN 55011 Class A Noise immunity ................. in compliance with DIN EN 61326-1 Laser Bench Rack/Rack 3F: Environment for use ......................Closed rooms Protection class .............................
  • Page 35 INSTRUMENT DESCRIPTION DirectFRAP Technical Data Carl Zeiss HXP 120: Environment for use ......................Closed rooms Protection class ............................I Ingress protection rating ........................IP 20 Overvoltage category ..........................II Line voltage range ..................100 V AC to 240 V AC ±10 % ....................
  • Page 36 INSTRUMENT DESCRIPTION Carl Zeiss Technical Data DirectFRAP SVB 1 (signal distribution box) Line voltage ......................100 V to 240 V ±10 % Change of line voltage is not required! Line frequency ......................... 50/60 Hz, Max. power consumption ......................20 VA Fuses 2x T 1.0 A/H...

  • Page 37: Start-Up

    The laser protection officer should be informed before the DirectFRAP imaging system is operated for the first time. The DirectFRAP imaging system has to be installed or retrofitted and adjusted in any case by the Carl Zeiss Microscopy GmbH Service or persons authorized by the manufacturer. If used as intended (see Section 2.1 on page 21), the DirectFRAP imaging system can be started up and...

  • Page 38: Power Requirements

    START-UP Carl Zeiss Installation Requirements DirectFRAP 3.1.2 Power Requirements Three wall outlets at a maximum distance of 1.5 m from the Laser Bench Tower or Rack are required to connect the system. Use the three multiple socket-outlets delivered to connect the system components (see Fig. 3-1).

  • Page 39: Ambient Conditions

    START-UP DirectFRAP Installation Requirements Carl Zeiss Fig. 3-2 Terminal assignments of the multipoint connectors for the DirectFRAP, Cell Observer SD (optional) and Laser TIRF slider (optional) combination systems 3.1.3 Ambient Conditions 1. Operation, specified power T = 22 °C ±3 °C without interruption (24 h per day,...

  • Page 40: Requirements For The Microscope Stand

    Axio Observer stand having an order number other than that indicated in the present manual, or without the safety kit for DirectFRAP, is not in accordance with the intended use. In particular, Carl Zeiss Microscopy GmbH cannot guarantee laser safety in such a case.

  • Page 41: Connecting The Fiber Oscillator

    START-UP DirectFRAP Connecting the Fiber Oscillator Carl Zeiss Connecting the Fiber Oscillator The fiber oscillator makes the optical fiber oscillate, providing homogeneous bleaching or photomanipulating of the ROI. • Place the fiber oscillator separately from the imaging system for laser manipulation, e.g. on the floor; it must not stand on the vibration-damped table.

  • Page 42: Inserting The Laser Manipulation Slider

    START-UP Carl Zeiss Inserting the Laser Manipulation Slider DirectFRAP Inserting the Laser Manipulation Slider • Carefully push the laser manipulation slider (Fig. 3-4/1) into the laser port of the microscope until it clicks into place (shortly before the mechanical limit stop is reached).

  • Page 43: Operation

    Carl Zeiss OPERATION The DirectFRAP imaging system is a class 3B laser system and labeled accordingly. CAUTION: This risk category includes medium-power lasers. Ensure that you are not exposed to the radiation of such lasers. In particular never look into the laser beam! Control and Functional Elements 4.1.1...

  • Page 44: Laser Bench Tower

    OPERATION Carl Zeiss Control and Functional Elements DirectFRAP 4.1.2 Laser Bench Tower Power input Power input Toggle switch (Tower O / I) Key switch (Laser ON / OFF) Power output Fig. 4-2 Control and functional elements of the Laser Bench Tower 4.1.3...

  • Page 45: Switching On

    OPERATION DirectFRAP Switching on Carl Zeiss Switching on If the laser manipulation slider is not yet located in the laser port of the microscope, insert it according to Section 3.5 (see page 42). • Visual inspection before switching on: Ensure that all safety covers (camera ports, fiber connections) are correctly mounted.

  • Page 46: Adjusting The Laser Beam With The Laser Manipulation Slider

    OPERATION Carl Zeiss Adjusting the Laser Beam with the Laser Manipulation Slider DirectFRAP Adjusting the Laser Beam with the Laser Manipulation Slider • Place the specimen slide used for the adjustment, with its cover glass facing downward, on the stage or put it into a mounting frame.

  • Page 47: Operation Via Zen Software (Blue Edition)

    The laser manipulation is controlled via the ZEN software (blue edition). For operating the software please see the ZEN operating manual (blue edition). DirectFRAP requires the fast acquisition module. The physiology module is recommended in addition. The planning and recording of the experiment is done using the software.
  • Page 48 83 (on) 83 (on) For combinations, a 100 % mirror inserted in the DirectFRAP is recommended for maximum dynamics to increase efficiency. The manipulation of Dendra with the laser line 488 is not possible in the "stand alone" DirectFRAP system.

  • Page 49: Care, Maintenance, Troubleshooting And Service

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE DirectFRAP Instrument Care Carl Zeiss CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE Instrument Care Care of the instrument by the user is restricted to the following operations: The instruments are not equipped with any special devices for protection against substances that are corrosive, potentially infectious, toxic or radioactive, or other substances that could be hazardous to health.

  • Page 50: Troubleshooting

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE Carl Zeiss Troubleshooting DirectFRAP Troubleshooting Problem Cause Troubleshooting No laser light Laser manipulation slider not inserted. Insert laser manipulation slider, see Section 3.5 on page 42. Reflected-light shutter is open. Close reflected-light shutter on microscope.

  • Page 51: Changing The Beam Combiner

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE DirectFRAP Changing the Beam Combiner Carl Zeiss Changing the Beam Combiner • Unscrew two screws (Fig. 5-1/1). • Push the cover of the beam combiner (Fig. 5-1/2, or Fig. 5-2/4) at an angle towards the front and remove it.

  • Page 52: Changing The Fuses

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE Carl Zeiss Changing the Fuses DirectFRAP Changing the Fuses Disconnect the power plug in any case before replacing the fuses. Axio Observer, HBO, XBO, HXP, Tempmodule S1: see separate operating manual. 5.5.1 Signal Distribution Box The fuse compartment is combined with the coupler plug and contains two fuses.

  • Page 53: Laser Bench Tower Coupling Module

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE DirectFRAP Changing the Fuses Carl Zeiss 5.5.3 Laser Bench Tower Coupling Module • Switch off the instrument by means of the power switch. • Disconnect the power plug. • Pull out fuse carrier (Fig. 5-6/1 and 3).

  • Page 54: Mounting The Camera

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE Carl Zeiss Mounting the Camera DirectFRAP Mounting the Camera The camera is connected to the sideport of the microscope using the camera adapter. To ensure safe operation of the laser the camera adapter is fitted with two reflected light barriers which close the laser shutter when no camera (Fig.

  • Page 55: Connect Hxp 120 Illumination Unit

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE DirectFRAP Connect HXP 120 Illumination Unit Carl Zeiss Connect HXP 120 Illumination Unit The HXP 120 illumination unit is connected by means of a coupling adapter, in which the optical waveguide is firmly secured and aligned, to the reflected light illumination connector of the microscope.

  • Page 56: Inserting Filters In The Excitation Filter Wheel 8X Mot

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE Carl Zeiss Inserting Filters in the Excitation Filter Wheel 8x mot. DirectFRAP Inserting Filters in the Excitation Filter Wheel 8x mot. • Tighten both screws (Fig. 5-9/1) on the excitation filter knob by hand. • Open the top side (Fig. 5-9/2) and put the excitation filter wheel (Fig.

  • Page 57: Mounting The Excitation Filter Wheel 8X Mot

    CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE DirectFRAP Mounting the Excitation Filter Wheel 8x mot. Carl Zeiss Mounting the Excitation Filter Wheel 8x mot. • Remove the cover of the reflected light mount (Fig. 5-11/1) of the microscope. • Insert the filter wheel with the dovetail mount (Fig.

  • Page 58: Service Request

    Any interventions to mechanical, optical and electronic components inside the device as well as work on the electrical system of the DirectFRAP imaging system for laser manipulation may be performed only by the Carl Zeiss Microscopy GmbH Service or by explicitly authorized specialist staff.

  • Page 59: Appendix

    APPENDIX DirectFRAP List of Abbreviations Carl Zeiss APPENDIX List of Abbreviations AOTF Acousto-Optical Tunable Filter Canadian Standards Association Deutsches Institut für Normung (German Standards Institute) European standard Electromagnetic Compatibility FRAP Fluorescence Recovery After Photobleaching Mercury-vapor short-arc lamp for fluorescence International Electrotechnical Commission...

  • Page 60: Index

    Laser Bench Rack ..........................44 Laser Bench Tower ...........................44 Control and functional elements ......................43 Diaphragm size ...........................30 Dimensions ............................33 DirectFRAP imaging system ......................21, 43 Experiment ............................47 Fiber oscillator .............................41 Functional principle ..........................22 Inserting filters in the excitation filter wheel ..................56 Installation notes ..........................
  • Page 61 APPENDIX DirectFRAP Index Carl Zeiss Maintenance ............................49 Microscope stand ..........................40 Mounting the camera ......................... 54 Mounting the excitation filter wheel ....................57 Notes on warranty ..........................20 Objective magnification ........................30 Operation ..........................33, 43, 47 Optical fiber ............................31 Power requirements ..........................

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