Leica EM FC7 Manual
Contents
1. Grip the assembly on the centre to retract the forceps Due to the backlash of the bearings the grid will be moved towards the knife edge if the assembly will be retracted by gripping on a different area of the assembly e g at the end of the Z movement assembly Avoid turning the micrometer gauges once the forceps is retracted The preset position would be changed thus the knife edge can be damaged if repositioning of the forceps is careless performed Start sectioning using the discharge mode of the Crion see chapter 5 6 When it is time to place the section ribbon on the grid switch off the motor of the specimen arm and the discharge function of the Crion For convenience both functions can be switched of by the footswitch simultaneously if Bind Motor to Crion pedals is enabled Page 43 Operating Manual Leica EM FC7 01 10 Page 44 Operating Manual Leica EM FC7 01 10 Slightly lift up the ribbon with the eyelash and advance the forceps to its pre set position Attach the ribbon on the grid using the charge mode of the Crion operated by the footswitch discharge For further section collection change the E W position of the grid and repeat as above described If you want to place the grid in the grid box move the grid box holder aside with tweezers and retract the forceps holding the grid Turn the forceps until the grid points downwards Make sure the grid does not interfere with parts of the2. Leica EM FC7 Operating Manual Version 162220032 01 2010 Living up to Life feica MICROSYSTEMS Important Note Leica reserves the right to change technical specifications as well as manufacturing processes without prior notice Only in this way is it possible to continuously improve the technology and manufacturing techniques used to provide our customers with excellent products Any copyrights of this document are retained by Leica Mikrosysteme GmbH Vienna Any reproduction of text and illustrations or any parts thereof by printing photocopy ing or other methods including electronic systems and media requires express prior permission in writing Issued by Leica Mikrosysteme GmbH Hernalser Hauptstrasse 219 A 1170 Vienna Operating Manual Leica EM FC7 Serial Number Date of purchase For the instrument serial number please refer to the name type label on the back of the instrument Please read this instruction manual carefully before operating the instrument Foreword This Technical Documentation is intended to provide essential information about the proper operation of the Leica EM FC7 Cryo chamber This user manual describes commissioning of the Leica EM FC7 Cryo chamber phased testing and adjustment of all components and movement sequences and restoring the basic functionality Service and operating staff must familiarize themselves with all components of the system before commissioning Part
3. Pull back grid holder Flip over bridge to the left Push forward grid holder until the grid is placed over the bridge Close section press by using a pre cooled pair of forceps Flatten sections under the press by pushing on the small indentation in the centre with a small black lever or a pair of forceps Apply pressure perpendicular to the surface to avoid sideways movement of the press Page 49 Operating Manual Leica EM FC7 01 10 Flip over section press to the right Release grid from the holder by pushing on the small indentation at the rear end with a small black lever or a pair of forceps and retract grid holder Transfer flattened grid into the transfer container The transfer container can be removed using a pair of forceps The ceramic surfaces of the section press should be cleaned with 100 ethyl alcohol before use Page 50 Operating Manual Leica EM FC7 01 10 5 11 Refilling the Dewar The 25l Dewar holds enough LN for a 1 day operation It is recommended to keep the pump in the Dewar until the LN has been consumed The LN consumption of the pump when not in use is less than 1I per day Refilling can be done in 2 different ways 1 Lift the pump slightly and refill the Dewar using the Dewar Refilling System 2 Take out the pump from the Dewar The cold pump should always be kept in an upright position to avoid entrance of water and humid air into the valves Use the yellow
4. debris If this occurs just clean the ionizer tip with a fine brush with the device switched off The electrode tip carries high voltage and must not be touched or allowed to touch any parts of the cryo ultramicrotome while operation Switch off the antistatic device prior to any manipulation in the chamber Never heat the chamber with the electrode inside Page 34 Operating Manual Leica EM FC7 01 10 5 7 Advanced touch screen control unit to operate the EM FC7 GD GID D 100 500 300 100 SPEEDmms FEEDnm Standard Mode Mr gt gt O 9 HEAT START 8 fal 5 00 100 et gt we gt Page 35 Operating Manual Leica EM FC7 01 10 1 LED chamber illumination Touch to switch on or off When activated brightness can be controlled with buttons 2 Integrated ioniser control Touch to switch on or off if EM Crion with discharge mode only is used When activated intensity can be controlled with buttons 3 Three individual readings for the actual temperature of specimen knife and gas Only the actual and not the set temperatures are shown Touch one two or all three buttons to activate setting mode When activated the temperatures can be set A number pad appears on the screen The desired temperature can be keyed in and must be confirmed with OK If only one button was activated e g knife only the temperat
5. 21 Operating Manual Leica EM FC7 01 10 4 4 Mounting the EM FC7 onto the Ultracut UC7 Make sure that the clamping lever of the knife support and the cryochamber are opened Move the knife stage of the EM UC7 into the E W centre position using the FC Mount button of controller or with the E W control wheel Page 22 Operating Manual Leica EM FC7 01 10 Take FC7 chamber with both hands and place it above the first indentation lines Push aside the chassis of the FC7 the groove on the chassis of the cryochamber should be on the right side of the line Slide on the chassis of the Ultracut UC7 until the second indentation line Turn the hand wheel of the Ultracut UC7 to move the specimen arm to the lowest position Guide the cryo specimen arm into the specimen arm of the Ultracut UC7 Slide the FC7 cryochamber into the chassis until you reach the end stop Lock the cryo specimen arm of the FC7 cryochamber to the specimen arm of the Ultracut UC7 by turning the locking screw either manually or using the 3 mm Allen key Observe the gap between cryo specimen arm and cryochamber 2 red arrows in The distance of the gap should be approx 0 5 mm Page 23 Operating Manual Leica EM FC7 01 10 Adjusting the distance between specimen arm and cryochamber should be done as follows Press the cryochamber to the right side and north against the UC7 or UC6 base plate red arrows The distance will be adjusted by turning t
6. protective cap immediately to close the lower end of the pump Before inserting the pump again dry pump and take off yellow protective cap For ease of operation one can use a second Dewar and place the pump immediately into the new full Dewar Taking out the pump from the Dewar if the pump is not in use for a longer time The cold pump should always be kept in an upright position to avoid entrance of water and humid air into the valves Use the yellow protective cap and the storing bracket that is provided with the pump Before removing the pump from the Dewar switch off pump and wait for a few minutes The LN hose must be flexible enough to be moved Page 51 Operating Manual Leica EM FC7 01 10 5 12 Finishing work with the EM FC7 Close down and bake out Pressing the Heat button will activate the automatic bake out of the chamber The FC7 cryochamber will now heat up to 110 C The heating cycle will be switched off automatically and the instruments will go back to room temperature During the heating time approximately 90 minutes the FC7 must be connected to the mains The heating cycle can be stopped and started again any time by pressing the Heat button e take out all specimens e take out diamond knives e take out antistatic electrode Before heating the chamber to avoid any damage to components and knives e unlock torque limited screw holding the cryo specimen holder This will prevent mechanic
7. side protection which are open at the top and at the bottom Only use boots if you have loose not narrow trousers coming outside the boots d and completely covering the gap Wear only open slip on sandals f in the lab no walking shoes or court shoes Always wear cuffless trousers if you wear slip on sandals Never wear protective gloves when pouring LN or when putting the Dewar head on the Dewar vessel Just use an open flannel cloth h to protect your hands from the cold Gloves should be used only to grasp dry cold parts They are unsuitable for LN work Page 7 Operating Manual Leica EM FC7 01 10 Only use metal Dewars specifically designated for storage of LN since only containers of this kind exclude risks during storage For routine cryopreparation metal troughs 1 cm styrofoam insulation styrofoam containers or plastic troughs are eminently suitable and ensure low risk cryopreparation Check the evaporation rate of your metal Dewar regularly every three months and compare these rates with the rate given by the manufacturer The evaporation rate of an undamaged metal Dewar should be well below 1 litre of LN per day Defective Dewar vessels with higher evaporation rates are a safety risk and should be taken out of work or repaired Do not leave LN standing in open vessels where it can exchange with the room atmosphere The boiling point of LN 196 C is lower than liquid oxygen s boiling point 183 C When the exchange
8. torque limited screw a using the 3 mm Allen key If the cryo holder is locked in the 0 position initially subsequent reading of the next position turned 90 for trimming will be easy 5 2 Inserting knives into the knife holder Page 30 Operating Manual Leica EM FC7 01 10 Set the desired clearance angle typically 6 Insert the desired knives e g a diamond trimming tool and a diamond 35 cryo immuno knife for Tokuyasu technique specimens Lock the knife by turning the locking screw clockwise Clearance angle setting and locking unlocking of knives can also be done with the knife holder in the cooled cryochamber 5 3 Inserting the knife holder The knife plate in the cryochamber carries a pin red arrow that corresponds to the hole in the knife holder red arrow Insert the rotating knife holder containing knives into the chamber The rotating knife holder is now in the loading position It can only be inserted in this position In the loading position the knife holder cannot be locked but is held in position by a magnet Page 31 Operating Manual Leica EM FC7 01 10 5 4 Inserting the specimen delicate knife edge Page 32 Operating Manual Leica EM FC7 01 10 Once the cryochamber has cooled down to the desired temperature e g 80 C for trimming of Tokuyasu technique specimens insert the specimen pin with the specimen Make sure the knife holder is in loading positi
9. al damage of specimen holder and locking screw due to heat expansion e unlock knife holder This will prevent mechanical damage of the locking screw due to heat expansion e move stereomicroscope with stereo carrier to the side This will prevent steam condensing water droplets on optical components 5 13 Maintenance of the valves VALVE 2 VALVE 1 Page 52 Operating Manual Leica EM FC7 01 10 Humidity in the valves can cause freezing and sticking of the valves during operation To prevent this both valves in the block can be easily removed and cleaned with ethanol Remove both steel valves with a screw driver The 2nd steel valve is located underneath the 1st valve Before inserting the valves again check that the ball inside the valves is free 6 Technical specifications Chamber illumination Top light Back light Spot light Cutting transmission Specimen advance Reserve warning Cutting speed Section thickness Return speeds Coarse knife movements N S Step control E W movement Knife holder Temperature range Temperature working range Temperature difference setting knife sample High gas flow advanced controller Automatic rapid cooling High gas flow advanced controller Temperature memories Page 53 Operating Manual Leica EM FC7 01 10 LED brightness controllable max 2150 lx LED brightness controllable max 8250 lx LED brightness controllable max 3450 Ix LED max 8600 Ix vibration dec
10. charge and charge mode or discharge only connecting cables Optionally available accessories Micromanipulator for frozen hydrated sectioning Set of cryotools Dewar Refilling System Cryo Mesacut Mirror Trimming Tool Different Flat Specimen Holders Page 17 Operating Manual Leica EM FC7 01 10 4 2 Preparation of the ultramicrotome UC7 or UC6 Page 18 Operating Manual Leica EM FC7 01 10 Remove armrest knife support and segment arc from the Ultracut EM UC7 or UC6 i Switch on the Ultracut Ultracut after removal of armrest knife support and segment arc Activate the FC Mount Function of the Ultracut by pushing the FC Mount button of the controller With the FC Mount Function of the Ultracut UC6 or UC7 the knife stage will be centered in the E W direction and to the outmost south direction automatically The backlight will switch off the toplight will switch on This will facilitate mounting the EM FC7 4 3 Unpacking the cryochamber EM FC7 Open box and lift out EM FC7 cryochamber and LN hose Page 19 Operating Manual Leica EM FC7 01 10 Remove LN hose and take away foam packaging Page 20 Operating Manual Leica EM FC7 01 10 Before mounting the EM FC7 on the Ultracut remove the transport locking screws on rear side and bottom knife plate of the cryochamber Keep the transport locking screws in the case for accessories for potential future shipping of the cryochamber Page
11. dling or to natural wear over many years of use Always keep the working area well ventilated Bring objects at room temperature carefully into contact with LN Initially an insulating gas layer is formed preventing a large transfer of heat During this initial period little LN evaporates However once the object has cooled down there may occur unexpected strong boiling and spurting of LN In the case of burns from LN splashes rinse the affected skin immediately with plenty of water at hand temperature For serious burns arrange for a skin specialist to see them at once In the case of LN affecting the eyes rinse immediately with water at hand temperature and arrange for an eye specialist to see it at once Never use glass Dewar vessels in the lab especially glass Dewars larger than 2 litres capacity without complete metal envelope Glass Dewars often burst for no obvious reason or due to unintentional mishandling e g contact with metal instruments etc Never work without open protective glasses when using LN in a glass Dewar Page 6 Operating Manual Leica EM FC7 01 10 Fig 1 1 When working with LN avoid protective glasses a boots c walking shoes e and protective gloves g out of which the LN cannot easily escape if entered LN splashing into the closed protective glasses a open boots c shoes e or protective gloves g evaporates suddenly and can cause serious burns Always use protective glasses b with
12. e Led re die E ee dag l Ae ve Paigpn Ven MEy prim PA ei OSMO connect cable to earth in the case the PE ground through the mains cord is not properly functioning remote control cable connect the other end to the rear side of the controller power cable 4 8 Connecting the EM CRION with discharge and charge mode Please see Leica EM Crion manual as well earth of the electrode electrode connect cable to earth in case the PE ground through the mains cord is not properly functioning remote control cable connect the other end to the rear side of the controller discharge to footswitch charge to footswitch power cable Page 26 Operating Manual Leica EM FC7 01 10 Advanced touch screen control unit A 1 0mm s 1 0mmis 3 0mmis 100mmis 50nm 7Onm 180nm 1 00pm Basic touch screen control unit Page 27 Operating Manual Leica EM FC7 01 10 In case a Leica EM UC7 instrument table is available place the power supply onto the shelf beneath the table board EM Crion with discharge and charge mode is operated by footswitches except setting the intensity The EM Crion with discharge function only is operated by switching on off with the button on the screen of the controller it can be switched on off if a footswitch is connected on the controller and Bind Motor to EM Crion is enabled see EM UC7 manual As soon the Crion is switched on a operating field
13. e LN in well ventilated areas Treat it with respect Storage For reasons mentioned above do not store full LN Dewars in confined spaces Burns LN gt boils at 196 C It is extremely cold and can cause serious burns Please read the safety instructions provided with all Leica products for the correct handling of liquid nitrogen Page 9 Operating Manual Leica EM FC7 01 10 1 1 Symbols and abbreviations Caution This symbol alerts the user to important information which may endanger staff or result in damage to the system if it is ignored Note This symbol indicates further information relating to a previous explanation which does not have a safety critical function However it is important to observe this information to ensure that the system functions optimally 1 2 Abbreviations CU controlunit n i not illustrated E W East West LN Liquid Nitrogen N S North South RC Rapid Cooling RT room temperature Page 10 Operating Manual Leica EM FC7 01 10 2 Identification 2 1 Product Leica EM FC7 Cryo chamber 2 2 Manufactured by Leica Mikrosysteme GmbH Hernalser Hauptstrasse 219 A 1170 Vienna Tel 43 1 488 99 0 Fax 43 1 488 99 350 Internet http www leica microsystems com Page 11 Operating Manual Leica EM FC7 01 10 3 Introduction 3 1 Overview The Leica EM FC7 is the low temperature sectioning system for the Leica Ultracut UC7 and UC6 ultramicrotome It is used f
14. e touched or allowed to touch any parts of the cryo ultramicrotome while operation Switch off the antistatic device prior to any manipulation in the chamber Never heat the chamber with the electrode inside Page 33 Operating Manual Leica EM FC7 01 10 5 6 Using the EM Crion with discharge and charge mode EM Crion with discharge and charge mode is operated by footswitches except setting the intensity For trimming For perfect ribbons of sections a perfect trimmed sample is mandatory It is recommended to use the discharge mode for trimming because it eliminates the sticking of the chips on the specimen and on the blade Set the intensity for trimming at the full voltage all 10 indication marks are illuminated Press the discharge footswitch to enable the antistatic performance or the electrode For sectioning Set the intensity to 5 indication marks If the sections tend to stick on the knife edge increase the intensity step by step until the sections begin to float in a nice ribbon over the knife edge If the sections begin to lift up decrease the intensity step by step Press the discharge footswitch to enable the antistatic performance or the electrode For placing the section ribbon on the grid The ribbon is attached to the grid by pressing the charge footswitch Thus the specimen adheres to the grid without the need of a section press The electrode will lose its power if its metal tip becomes covered with ice or
15. h O Fahrenheit Show Cutting Animations O Kelvin Show Info Messages Click OK to save or Cancel to discard settings Show Confirm Messages Play Sound on Button Click sona Stage Feed Minimum 2500 o Edit sample Lever Cutting Settings Lever 7 FC Mode 2 Following settings are Standard Mode stored for the current sample and can be modified using the speed feed knobs and the controls on main screen High Gas Flow Click DK t ioni speed Feed Presets Cancel to SE SL Return Speed settings Garen FC temperature settings lz ion can be modified on the Antistatic main screen Counter values Autotrim values Cancel Page 37 Operating Manual Leica EM FC7 01 10 1 Temperature Unit After pushing the menu button on the main screen the temperature unit can be selected in the Edit User menu 2 FC Mode The FC7 chamber is equipped to toggle between three different modes Push the menu button on the main screen and select the Edit or Add New sample menu o Standard mode is preferable used for regular sectioning e g Tokuyasu or polymer sectioning at dry conditions o High Gas Flow is preferable used to increase GN gas flow below a temperature of 140 C in order to reduce ice contaminations o Wet Sectioning mode is used for polymer sectioning at wet conditions using DMSO or similar liquids in order to set the temperature of knife at a maximum of 40 C and
16. h screen control unit 3 2 Instrument Function Operation of the EM FC7 is performed via the control Unit of the Ultracut UC7 Before quickly cooling down for trimming and sectioning the cryochamber has to be unpacked mounted on the EM UC7 and connected to the Ultracut UC7 CU As the cryochamber is mounted on the chassis of the ultramicrotome it is isolated from external vibrations The contact free through the wall specimen arm provides optimum temperature and mechanical stability After connection of the LN hose to the LN filled Dewar via the automatic pumping system the pump ensures a stable low temperature environment around the knife specimen area by pumping LN into a tank in the EM FC7 wall This LN is then heated and the evaporating gas is used to cool down chamber specimen and knife Temperature stability of specimen knife and gas in the chamber are individually controlled by regulators close to knife and specimen The desired temperature can be set changed and stored by the operator via the control unit while the actual temperatures are displayed on the CU After finishing cryosectioning a bake out function allows the cryochamber to heat up to 110 C before returning to room temperature RT automatically Page 13 Operating Manual Leica EM FC7 01 10 3 3 Advanced touch screen control unit After connecting and switching on the EM FC7 and the EM CRION to Advanced touch screen control unit of the Ultramicrotome UC7 the to
17. he adjusting screw on the front side of the FC7 cryochamber using the 2 mm Allen key For accurate setting the 0 5mm spacer transparent plastic foil can be used Lock the FC7 cryochamber to the chassis of the Ultracut UC7 by turning the lever a clockwise until its end stop position Di Make sure the FC7 is firmly connected by gently pulling in the south direction To increase the clamping remove lever a by unscrewing slot screw Reposition the lever an eighth counter clockwise on the clamping shaft and lock FC7 Repeat procedure until FC7 cannot be removed by gently pulling in south direction Fix lever by tightening the slot screw Push the knife plate of the EM FC7 until its north end stop position Lock the knife plate to the knife stage of the Ultracut UC7 or UC6 by turning the lever b clockwise Move the cryo specimen arm up and down by turning the hand wheel to make sure that there is no contact to the cryo chamber Make sure the cryochamber is levelled out with a spirit level placed on the top plate of the cryochamber see operating manual of the EM UC7 as well 4 5 Connecting the Cryochamber Before connecting the power supply to the main following main supply must be assured 100 240 VAC 50 60 Hz The instrument is equipped with protected ground Before connecting it to the local electrical supply make sure that the mains has the required ground and that the instrument is connected to it according t
18. icular attention must be paid to the aspect of safety This user manual must be retained for future reference Texts schedules and tables may not be copied reproduced or divulged to third parties without our express consent It should be noted that this Technical Documentation does not constitute a part of any existing prior agreement or covenant or legal relationship All obligations are derived from the purchase agreement which is also the sole document of record regarding the terms of the warranty Contractual provisions are not affected by the Technical Documentation The documentation issued by the respective suppliers shall apply in addition to this manufacturer s documentation In addition all generally applicable legal and otherwise binding regulations for preventing accidents and protecting the environment must be observed and communicated Page 2 Operating Manual Leica EM FC7 01 10 Table of contents i Safety NCE OCC OI Rail 6 1 1 Symbols and abbellire 10 1 2 PODISVIAUOIS sorna on idas 10 2 Ted E 11 2 1 PIOGUCE EE 11 2 2 Manufactured iii 11 gie Introduction a nai 12 3 1 A A re iO CO A 12 3 2 PTS CUTTS UN COI EE 18 3 3 Advanced touch screen control UA orcos 14 3 4 Basic touch screen control UN LEET 15 4 Installation and Te iniziata 16 4 1 A A i lo 16 4 2 Preparation of the ultramicrotome UC7 or UCp 18 4 3 Unpacking the cryochamber EM FOT 19 4 4 Mounting the EM FC7 onto the Ultracut UC 22 4 5 Co
19. id can be performed manually prior to sectioning to prevent possible influence of the grid on the ionizer The Leica EM CRION is used in discharge mode in order to reduce electrostatic charging while sectioning When it is time to place the section ribbon on the grid the user can quickly return to the pre set grid position The ribbon is then attached to the grid using the charge mode of the LEICA EM CRION which is operated by a footswitch Thus the specimen adheres to the grid without the need of a section press As a grid box can be placed near the knife the micromanipulator can be used to easily place the grid into its storage position Place the grid box holder with its magnet onto the knife holder Insert the grid box in the holder Page 41 Operating Manual Leica EM FC7 01 10 Page 42 Operating Manual Leica EM FC7 01 10 Pick up the grid with the forceps For accurate manipulation hold the forceps close to the tips Insert the forceps in its holder of the micromanipulator and slide it towards the end stop position With the micrometer gauges set the grid slightly below the knife edge with the z movement E W position of the sample and the N S position close to the knife Make sure the shaft of the micrometer gauges are in contact with the magnetic end stop N S movement E W movement Z movement To prevent possible influence of the grid on the onizer retract the forceps by pulling back the assembly
20. knife holder It might be necessary to move the forceps in a proper E W position as well Lift up the forceps either by using the micrometer gauges or manually by gripping the z direction lever Adjust the grid box using a pair of tweezers and the grid with the y and x movement of the manipulator in order to position the grid above an empty recess of the grid box Lower the forceps slowly until the grid is placed in its position of the grid box Open the forceps to release the grid To use a new grid take out the forceps and warm it up as long as the forceps is dry To speed up the process a second forceps is recommended Or another possibility is to load the second grid holder with empty grids and place it in the chamber If a new grid has to be inserted into the forceps exchange the grid boxes and pick up the empty grid using the micrometer gauges of the micromanipulator In this case the forceps can remain in the cryochamber and will not ice up if moved out of the chamber Page 45 Operating Manual Leica EM FC7 01 10 5 10 Set of Cryotools The Set of cryotools is used for handling frozen hydrated sections Tokuyasu sections as well as collecting polymer sections 5 10 1 Unpacking The set consists of 4 parts the base with a flip over bridge for preloading releasing of grids the spring loaded grid holder the section press and the transfer container 5 10 2 Device function and operation The base of the cryotoo
21. ls is mounted on the rotating knife holder with a locking screw A pre cooled grid is clamped in the spring loaded grid holder 2 The grid holder is shifted forward until the grid is close to the knife edge Cryosectioning is carried out Once the ribbon of frozen hydrated sections is on the grid the holder with the grid is pulled back in south direction The grid is then placed onto the flip over bridge 3 and sections are pressed with the section press 4 After flattening the sections on the grid the grid is transferred to a transfer container 5 Page 46 Operating Manual Leica EM FC7 01 10 Insert cryo knife into knife holder Mount base of cryotools on rotating knife holder and lock Insert grid holder and transfer container into base of cryotools Flip section press over to the right Page 47 Operating Manual Leica EM FC7 01 10 Place grid on flip over bridge Open grid holder by pushing on the small indentation at the rear end with a small black lever or a pair of forceps Move gird holder towards the grid and clamp it by releasing the clamping jaw Pull back grid holder towards you and flip bridge to the right using a pair of forceps Push grid holder forwards until the grid has reached the desired distance from the knife edge Take extra care not to touch the delicate diamond knife edge Page 48 Operating Manual Leica EM FC7 01 10 Start cryo sectioning and collect the ribbon on the grid
22. nnecting the Een E le 24 4 6 Connecting the LN hose of the pump 25 4 7 Connecting the antistatic device EM CRION with discharge mode 26 4 8 Connecting the EM CRION with discharge and charge mode 26 4 9 Installing the Micromanipulator E 28 4 10 Repacking to prevent damage during transportation nsenseeeneneneeeneenereenresrrrrrrenrnrnnrenne 29 4 11 Storage location for the Imstruchons e 29 5 ee E d Lu EEN 30 5 1 Inserting the cryo specimen holder bk 30 02 Inserting knives into the knife holder 30 9 3 inserting the knife noO anale 31 5 4 Inserting the specimen arios paladar abia 32 Di Using the antistatic device EM Crion with discharge Mode 33 5 6 Using the EM Crion with discharge and charge mode 34 5 7 Advanced touch screen control unit to operate the EM FC7 35 5 8 Basic touch screen control unit to operate the EM FC 39 5 9 Operating the micromanipulator iii 41 SIS oO ee Lei EE 46 5 10 1 WMC I e BE 46 5 10 2 Device function and operaton i 46 5 11 Refilling the Re ET 51 5 12 Finishing work with the UNE 52 5 13 Maintenance of the valves erosiones toto 52 6 Technical specificatioliS airline diari 53 Page 4 Operating Manual Leica EM FC7 01 10 Page 5 Operating Manual Leica EM FC7 01 10 1 Safety Introduction Liquid Nitrogen LN2 When working with liquid nitrogen LN2 please bea
23. o the local regulations Unplug the instrument before installing or changing fuses In case a Leica EM UC7 instrument table is available place the power supply onto the shelf beneath the table board Connect the power supply to the mains Connect hot plug of the power supply to the multiway connector on the rear side of the ultramicrotome b Connect hot plug of cryochamber d Connect hot plug of pump a Multiway connector with hot plugs for pump a power supply b control unit c and cryochamber d Page 24 Operating Manual Leica EM FC7 01 10 4 6 Connecting the LN hose of the pump precautions Page 25 Operating Manual Leica EM FC7 01 10 Connect the LN hose with its threaded connection to the FC7 cryochamber Connect the LN hose with its threaded connection to the FC7 to the pump Before inserting the connection open the safety shutter a The clean Dewar vessel has to be filled with LN according to the safety precautions Remove the yellow protective cup from the lower end of the pump Slowly lower the pump into the Dewar Hold the pump for a while until the strong boiling decreases and carefully lower the pump until it is completely immersed The clean Dewar vessel has to be filled with LN according to the safety 4 7 Connecting the antistatic device EM CRION with discharge mode Please see Leica EM Crion manual as well earth of the electrode Tom CEA E Een Ae geng P electrod
24. on the screen of the control unit appears The intensity of the ioniser is controlled by pushing buttons on the ANTISTATIC field Place the antistatic electrode with its holder into its position on the cryochamber 4 9 Installing the Micromanipulator Unpack the micromanipulator supporting plate pair of forceps end stop grid box holder and two grid boxes Allen key Remove the top plate of the cryochamber by unscrewing the screws and exchange it with the supporting plate of the micromanipulator screw in the end stop plug in the ground cable locating plates for the electrode holder right plate for left handed operation of the eyelash Page 28 Operating Manual Leica EM FC7 01 10 4 10 Repacking to prevent damage during transportation The Leica EM FC7 must not be transported unless it is in its original packaging if it is not the system may be damaged If the system needs to be moved again disassemble and pack it away by following the instructions listed in Unpacking the cryochamber 4 3 in the reverse order 4 11 Storage location for the instructions The user manual and associated supplementary documentation e g manual of the EM Crion must be kept close to the Leica EM FC7 for fast access Page 29 Operating Manual Leica EM FC7 01 10 5 Operation 5 1 Inserting the cryo specimen holder Insert the cryo specimen holder into the specimen arm Lock the cryo specimen holder with the
25. on to avoid touching the Lock the specimen pin in the cryo specimen holder Turn the small black lever in front clockwise while holding the cryo specimen holder in position with the small black lever in the back Rotate the knife holder into trimming position Lock the locking screw of the knife holder with the 3 mm Allen key Start trimming and sectioning To change from sectioning position 1 e g the trimming knife to sectioning position 2 e g the sectioning knife unlock the screw rotate knife holder and lock again 5 5 Using the antistatic device EM Crion with discharge mode For trimming For perfect ribbons of sections a perfect trimmed sample is mandatory It is recommended to use the antistatic device for trimming because it eliminates the sticking of the chips on the specimen and on the blade Set the intensity for trimming at the full voltage all 10 indication marks are illuminated For sectioning Set the intensity to 5 indication marks If the sections tend to stick on the knife edge increase the intensity step by step until the sections begin to float in a nice ribbon over the knife edge If the sections begin to lift up decrease the intensity step by step The electrode will lose its power if its metal tip becomes covered with ice or debris If this occurs just clean the ionizer tip with a fine brush with the antistatic device switched off The electrode tip carries high voltage and must not b
26. or semi and ultrathin cryosectioning of biological and industrial samples This includes Frozen Hydrated Sectioning FHS of vitrified specimens cryoprotected biological samples Tokuyasu technique and soft industrial specimens such as rubbers and polymers The Leica EM FC7 has been specifically designed for maximum ergonomics and ease of use lt has an integrated chamber illumination allows to use an integrated antistatic device and provides a wide temperature range from 15 C to 185 C for cryosectioning All controls are via the control unit CU of the ultramicrotome to save workspace on the table and provide maximum comfort and ergonomics for the operator The EM FC7 system consists of Cryochamber with integrated LED illumination Dewar vessel on mobile trolley Liquid nitrogen pump Power supply Connecting cables LN hose Accessories box containing 2 rotating knife holders 1 standard specimen holder specimen carriers special keys tools and forceps The complete working outfit contains in addition to the items mentioned above Flat specimen holder CRION antistatic device remote controlled This allows controlling the ioniser from either the Advanced touch screen controller or the 7 touch screen controller of the Ultracut UC7 Recommended accessories are Micromanipulator Filling System for LN Dewar Page 12 Operating Manual Leica EM FC7 01 10 Leica EM FC7 mounted on Ultracut UC7 with Advanced touc
27. oupled gravity stroke 200um 20um 0 05 100 mm s wheel controlled 0 15000nm wheel controlled 10 30 50mm s 10 mm stepping motor 0 1 15 um steps 10mm stepping motor for two 6 10 mm knives 110 C to 185 C 15 C to 185 C 40 C knife and 170 C sample lt 165 lt 140 C 4 advanced controller 3 basic controller EC Declaration MICROSYSTEMS EC Declaration of Conformity EG Konformit ts Erkl rung Declaration CE de Conformite We Wir Nous Leica Mikrosysteme GmbH Hernalser Hauptstrasse 219 A 1170 Wien Austria declare in exclusive responsibility that the product erkl ren in alleiniger Verantwortung dass das Produkt d clarons sous notre seule responsabilit que le produit Model LEICA EM FC7 Modell LEICA EM FC7 modele LEICA EM FC7 Type Typenbezeichnung type EM FC7 706030 656002 to which this declaration relates is in conformity with the following standards auf das sich diese Erkl rung bezieht mit den folgenden Normen bereinstimmt auquel se r f re cette d claration est conforme aux normes EN 61010 1 EN 61326 1 following the provisions of directive gem ss den Bestimmungen der Richtlinie conform ment aux dispositions de directive 2004 108 EC Electromagnetic compatibility Elektromagnetische Vertr glichkeit 2006 95 EC Low Voltage Equipment Niederspannungsrichtlinie 2006 42 EC Machinery Maschinen Merde Af Wien 25 June 2009 Dr Reinhard Lihl Entwicklung
28. r in mind LN is extremely cold It boils at 196 C Nitrogen gas GN escapes at very low temperature from the boiling LN Both LN and GN as well as cooled elements e g pipes valves hoses containers or stoppers can cause severe frost bite and burns to the skin and eyes When LN evaporates it expands in a ratio of 1 700 1 litre LN produces almost 1 m3 of GN Care should therefore be taken to ensure that when large quantities of nitrogen evaporate e g when transferring LN2 the room should always be well ventilated Removing LN waste dump LN into an outdoor pit or container filled with gravel where it will evaporate rapidly and safely GN gt is odourless and tasteless and will be inhaled like air GN is non toxic but a high GN content in the air gt 78 reduces the oxygen content lt 21 and produces immediate fainting and deep unconsciousness without any previous symptoms When there is doubt about the adequacy of ventilation use an oxygen analyser 0 to 25 scale to check for oxygen The content of oxygen must not drop below 18 If an unconscious person stays in a low oxygen environment then death may occur If breathing stops apply artificial respiration at once and notify doctor and ambulance immediately For the reasons given above never put LN Dewars in a closed storage room or chamber The evaporation rate from Dewar vessels can rise to several litres a day if they are defective due to improper han
29. sleiter R amp D Manager Chef du service d veloppement Leica Microsysteme GmbH Tel 43 1 48899 Hernalser Hauptstrasse 219 Fax 43 1 48899 350 A 1170 Wien www leica microsystems com OLOZ LO ZE00ZZZ9 UOISISA 194 INA 29197 enueyy Bunesado uonipa ou jo ou pio IN HOWE yl 29197 Jo sxuewape pauajsifau ale obo e9197 ay pue wO 0S 6688h E XC 6688Y Ep IAL ESNY Su 0211 GLZ essessidney JasjeuJay Hquo auejs som IN 29197 1U6uAdog MICROSYSTEMS www leica microsystems com
30. stored value The reading for the actual temperature shows the set value for approx 4 seconds To store a new value set temperature 3 and press one of the memory buttons An acoustic signal confirms the input 5 LN level indicator Indication of the filling status of the Dewar An acoustic warning signal will sound if the Dewar is almost empty Approx 1 h working time is left The acoustic signal can be switched off by touching the Dewar symbol 5 or the displayed hint box appears in the upper left corner on the screen Menu 6 LN stop Touch to switch on or off pump e g for refilling the Dewar it may be necessary to warm up the LN hose until it is flexible enough to me moved 7 One of four different command variables can be set The temperature will be controlled according the selected set point T Average an average of specimen sample and gas Tk Knife Ts Sample Tg Gas 8 Temperature Unit a choice of C F and K is available The selected unit will be displayed on the main screen Page 40 Operating Manual Leica EM FC7 01 10 5 9 Operating the micromanipulator The micromanipulator in conjunction with the EM CRION with discharge and charge mode is indented to use for section collection of frozen hydrated sections as well as Tokuyasu sections The micromanipulator allows the grid to be exactly positioned close to the knife edge using the micrometer gauges Once these positions are defined fast retraction of the gr
31. surfaces are extensive enough oxygen from the air will be taken up in exchange for nitrogen LN with high liquid oxygen content has a faintly bluish colour Concentrated liquid oxygen promotes vigorous burning Make sure that your Dewar vessel is filled only with LN Apply a note in the central distribution place stating clearly ONLY LIQUID NITROGEN or similar if different liquefied gases are delivered from there Check the colour of cryogen Bluish colour indicates the presence of a high percentage of liquid oxygen The concentration of liquid oxygen increases during long periods of storage as its boiling point 183 C is higher than the boiling point 196 C of LN Main supply must be assured 100 240 VAC 50 60 Hz The instruments are equipped with protected ground Before connecting it to the local electrical supply make sure that the mains has the required ground and that the instrument is connected to it according to the local regulations Unplug the instrument before installing or changing fuses Page 8 Operating Manual Leica EM FC7 01 10 HAZARD WARNING LIQUID NITROGEN CN Suffocation Any vessel containing LN is a potential hazard One litre LN produces 700 litres No gas N2 gas is odourless and tasteless Oxygen levels can quickly drop in confined spaces due to displacement of oxygen by No when using or dispensing large volumes of LN This can cause immediate fainting and unconsciousness Always us
32. the temperature of the sample minimum of 170 C To achieve the minimum temperature of the sample the specimen arm must be moved to its lowest position during the cooling process Thus parts of the cooling element of the sample holder are placed into LN which facilitates cooling down the sample at a minimum of 170 The temperature will be kept during the sectioning process Page 38 Operating Manual Leica EM FC7 01 10 5 8 Basic touch screen control unit to operate the EM FC7 Leica MM MICROSYSTEMS Menu Version 04 01 02 Bind Motor to EM Crion Factory Defaults Page 39 Operating Manual Leica EM FC7 01 10 1 LED chamber illumination Press to switch on or off 2 Start press to activate cooling of the chamber to the set temperature After Start has been pressed the name of the button changes to Heat Press Heat to activate automatic bake out of the chamber The heating process can be interrupted by pressing the Heat button again Stop Heat Due to the automatic bake out the controller can be switched off and the bake out process will continue to heat up the chamber to about 110 C and stops automatically at room temperature afterwards 3 Reading for the actual temperature which is selected in the menu as command variable set point This temperature can be set with buttons By pressing the displayed value the set temperature will be shown for about 4 seconds 4 Three memory buttons Press to activate the
33. uch screen appears as shown below j Standard Mode WW 8 gt ANTISTATIG e wer wg Page 14 Operating Manual Leica EM FC7 01 10 3 4 Basic touch screen control unit After connecting and switching on the EM FC7 and the EM CRION to the 7 touch sensitive control unit of the Ultracut UC7 the main screen of the controller appears as shown below Leica MM MICROSYSTEMS Cut ge 7 Start 100 50 _ 0 gt cre ES ES End 7Onm 500 Uni 500 4 II Page 15 Operating Manual Leica EM FC7 01 10 4 Installation and Setup 4 1 Packing list LEICA EM FC7 Low Temperature Sectioning System contains EM FC7 cryochamber Power supply connecting cables Pressure less automatic filling system Insulated threaded connecting hose Dewar for 25l LN with special flange and mobile trolley and pump storage holder The case for accessories contains Specimen carriers 50 pcs with concentric rings Specimen carriers 50 pcs with slot Allen key 3 mm Cryotool with M4 thread Collet specimen clamp Rotating knife holder 2pcs Preparation plate Allen key 2mm Special long forceps with insulation coating Black lever 2 pcs 0 5mm spacer transparent plastic foil Transparent cover used during the cooling down process The Complete Working Outfit contains in addition Page 16 Operating Manual Leica EM FC7 01 10 EM CRION antistatic device either with dis
34. ure of this button in this case the knife will be set If all three buttons were activated all three will be set together 4 Store function Touch to activate storing mode followed by one of the four memory fields 5 The set temperatures of specimen knife and gas are now stored in the desired memory field 5 Memory fields Touch to activate the pre stored values 6 Pump Touch to switch on or off pump e g for refilling the Dewar it may be necessary to warm up the LN hose until it is flexible enough to me moved 7 LN level indicator Indication of the filling status of the Dewar An acoustic warning signal will sound if the Dewar is almost empty Approx 1 h working time is left The acoustic signal can be switched off by touching the Dewar symbol 8 Start Touch to activate cooling of the chamber to the set temperature 9 Heat Touch to activate automatic bake out of the chamber The automatic safe guard will ask you to touch the button for more then 1 second to prevent activation by accident Due to the automatic bake out the controller can be switched off and the bake out process will continue to heat up the chamber to about 110 C and stops automatically at room temperature thereafter Page 36 Operating Manual Leica EM FC7 01 10 MENU gt User or Sample profile Edit userprofile Andi Enable Manual Cutting Temperature Unit f Andi E z Bind Motor to EM Crion Pedals Celsius 1 Enable Continuous Approac
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