Leica CryoJane User Manual
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Leica CryoJane User Manual

Leica CryoJane User Manual

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CryoJane
User Manual
Customer Service 1-800-237-2772
(In continental US only)
Customer Service 1-201-343-1313
(International)

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Summary of Contents for Leica CryoJane

  • Page 1 ® CryoJane User Manual Customer Service 1-800-237-2772 (In continental US only) Customer Service 1-201-343-1313 (International)
  • Page 2 Instrumedics grants the purchaser of the CryoJane system the rights to use the patented CryoJane (CFSA) materials and process. Patent # 4,545 831...
  • Page 3 CRYOJANE TAPE-TRANSFER STEP-BY-STEP INSTALLATION INSTRUCTIONS The following items are required for installation: Absolute Alcohol 1600 Watt Hair Dryer Gauze Gloves 02 Place ECU on top left 01 Remove release paper 03 Using absolute alcohol and gauze from ECU corner of cryostat clean chamber wall 04 Heat left wall for Mech &...
  • Page 4 Cryo-Vac Shown connected to ECU Cryo-Vac Shown *The standard CryoJane Mech installation is shown in Fig. 07. The Hacker/Bright OTF and Microm HM 560 cryostats require an S-Plate Mech and separate mounting plate for installation. S-Plate Mech for Hacker OTF and Microm HM 560 Cryostats Mounting Plate locks onto S-Plate Mech.

  • Page 5: Table Of Contents

    Table of Contents Instrumedics Frozen Section Process Overview …………… Freezing Process …………………………………………………………. 7 CryoJane Process ………………………………………………………… 7 CryoJane System Components …………………………………….. CryoJane ECU ……………………………………………………………. CryoJane Mech Components ………………………………………. CryoJane Operating Procedure …………………………………… 11 Background ………………………………………………………………. 11 Preparation ……………………………………………………………….. 11 Use ……………………………………………………………………….. 12 Steps for a Single Section …………………………………………...
  • Page 6 Troubleshooting Guide … … … … … … … … … … … … … … … 23 Troubleshooting the Mech and ECU …………………………………….. 23 CryoJane Process Troubleshooting ………………………………………. 25 Troubleshooting Finished Slides …………………………………………. 27 Oil Bath Accessories Kit Components …………………………..

  • Page 7: Instrumedics Frozen Section Process Overview

    3. The curing of the adhesive on the slide with an 8 millisecond ultraviolet flash, and the subsequent removal of the adhesive tape, leaving a still frozen section firmly adhered to the microscope slide. The CryoJane system is designed to permit the facile application of these methodologies, and can be adapted to virtually all model cryostats.

  • Page 8: Cryojane System Components

    CryoJane System Components FIGURE 1. CryoJane System Components 1. Mechanism (Mech). See CRYOJANE MECH COMPONENTS for details. 2. Electronics Control Unit (ECU). See CRYOJANE ECU for details. 3. ½ inch diameter Mech Cable (connected to the Mech) 4. Door Channel 5.

  • Page 9: Cryojane Ecu

    (see Cryo-Vac-Away User Manual) CryoJane ECU Figure 3. CryoJane ECU - front and right side Figure 2. CryoJane ECU - back 1. On/Off Switch 2. Pad LED 3. Power LED (or Bath LED if optional Oil Bath Accessory Kit is installed).

  • Page 10: Cryojane Mech Components

    CryoJane Mech Components FIGURE 4. CryoJane Mech Components 1. Blue Pad 2. Stainless Steel Spring Clip 3. Black Knob 4. Flash Tray 5. Lid...

  • Page 11: Cryojane Operating Procedure

    Pathologists and clinicians have been trained to recognize normal and abnormal structure against this background of degraded fine structure. In contrast, the CryoJane process is designed specifically to avoid melting and air drying and its associated artifacts. It uses a cold adhesive tape to support and capture the section during the cutting step.

  • Page 12: Use

    • The right LED should be red if it is labeled Power and green if it is labeled Oil Bath. Once a tissue specimen has been frozen and mounted in the microtome chuck, follow the procedure outlined below. Steps for a Single Section: 1.
  • Page 13 3. Holding the Slide at the frosted end, rapidly peel off the protective mylar film which covers the adhesive and discard it. Figure 7. Peeling mylar from adhesive coated slide. 4. Place the Coated Slide, adhesive-side- up, on the Blue Pad of the Mech by slipping the frosted end of the slide under the stainless steel Spring Clip.
  • Page 14 6. Hold the Tape up by the narrow tab with the adhesive side towards the block face. Position the Tape in front of the trimmed block so that the bottom edge of the exposed adhesive is aligned with the bottom edge of the block. Place the Tape on the block face making sure that the bottom wider tab of the Tape overhangs the knife.
  • Page 15 8. Turn the flywheel slowly and evenly to cut a section. If a sharp knife is used, the section captured on the Tape will be flat, intact, uncompressed and as thin as two microns. No need for a brush or an anti-roll device! Figure 12.
  • Page 16 Figure 14. Positioning the Tape on the Slide. 11. Use the cold Hand Roller to laminate the Tape to the Slide. Apply even pressure on the Roller and roll it over the section under the Tape two or three times Figure 15.
  • Page 17 14. Holding the frosted end, immediately remove the cured Slide from the Flash Tray. DO NOT TAKE THE SLIDE OUT OF THE CRYOSTAT AT THIS TIME AS THAT WILL CAUSE THE SECTION TO MELT. Figure 18. Removing the cured Slide. 15.

  • Page 18: Steps For Multiple Sections

    a) fixed in the Instrumedics Aqueous Fixative, b) freeze-dried or freeze-substituted in the cryostat before “anhydrous” fixation, Note: The section should be immersed in the fixative inside the cryostat. Figure 20. Technique for removing Tape from Fixation should then continue at room Slide.

  • Page 19: Summary Of The Instrumedics Frozen Sectioning Process

    19. Close the Lid and push and release the Black Knob to actuate the UV flash. Then lift the Lid, remove the Slide and carefully peel away each strip of Tape as described in Steps 15 and 16. Figure 22. Curing multiple sections. Summary of the Instrumedics Frozen Sectioning Process Snap-freezing and embedding.

  • Page 20: Special Fixation

    Special Fixation Protocols Aqueous Fixative: THIS FIXATION SHOULD BE USED FOR ALL HISTOLOGICAL STAINING OF CRYOJANE PREPARED SECTIONS. This fixative melts and fixes the tissue simultaneously (direct controlled “melting”). Ice crystal artifacts (holes) are masked. Nuclear morphology is usually excellent. Some cell components are lost and most of the red blood cells are lysed.

  • Page 21: Anhydrous" Fixation

    a) After the adhesive coating on slide has been polymerized, carefully peel off the pink Tape. b) To avoid uncontrolled melting and/or drying of the section, immediately bring the room temperature aqueous fixative into the cryostat and immerse the slide into the fixative.

  • Page 22: Cryojane Maintenance

    30 to 60 seconds. This transfer must be made within the cryostat. Following fixation, the slides are rinsed in water prior to staining. REFER TO REAGENT MSDS FOR SAFE HANDLING CryoJane Maintenance • NOTHING SHOULD BE PLACED OR STORED ON TOP OF THE ECU! Any spills which enter the ECU may severely damage the electronic components.

  • Page 23: Troubleshooting The Mech And Ecu

    • Liquid spills on the Mech should be wiped immediately. Electronic components in the Flash Lamp may short circuit if exposed to liquids. • DO NOT IMMERSE THE MECH IN ANY SOLUTIONS. • To remove the Mech from the wall for decontamination or servicing, simply unscrew the two screws shown below and then remove the Mech from the cryostat.
  • Page 24 Pad connector underneath the Mech. The temperature of the Blue Pad is too cold. Raise the cryostat temperature. Call Instrumedics’ 800 customer service The ECU is malfunctioning. number and arrange for servicing of the ECU (1-800-237-2772). DO NOT OPEN THE ECU! ♦...

  • Page 25: Cryojane Process Troubleshooting

    ECU (1-800-237-2772). DO NOT OPEN THE ECU! CryoJane Process Troubleshooting The CryoJane process will not produce the desired results if the cryostat and/or the microtome are malfunctioning. Both need to be serviced periodically. Check with the manufacturer/dealer.
  • Page 26 degrees. The Tape is defective. Use another Tape. ♦ The Tape is cut during sectioning. Cause Solution The block face is not flat. Align and trim the block to obtain a flat face. The adhesive on the Tape overhangs the block Position the Tape on the block so that no and snags on the knife.

  • Page 27: Troubleshooting Finished Slides

    Remove the Tape slowly and carefully, peeling The Tape is not being removed properly. it diagonally and downward to minimize the tension on the section. If using a disposable blade, shift to a different The knife is dull. portion of the blade’s edge or replace the blade.
  • Page 28 Slide. surfaces of the knife’s cutting edge before sectioning. ♦ The section did not transfer completely. See the CryoJane Process Troubleshooting Guide. ♦ There are holes in the nuclei and/or cytoplasm of the section. Cause Solution Transfer the frozen block from the freezing device rapidly to the cryostat to avoid thawing the tissue.
  • Page 29 Snap-freeze tissue at -60°C or colder. Using Instrumedics’ Power Jane, or Stand Alone Gentle Jane or Dewar Jane snap-freezers with The initial freezing was poor (slow), causing liquid nitrogen produces the best results. large ice crystals to grow. The section was Snap-freeze the thinnest practical specimen.

  • Page 30: Oil Bath Accessories Kit Components

    Oil Bath Accessory Kit Components FIGURE 28. Oil Bath Accessory Kit Components 1. Oil Bath 2. Oil 3. Oil Brush 4. Oil Bath Power Cable (not shown)

  • Page 31: Oil Bath Accessory Kit Operating Procedure

    Oil Bath Accessory Kit Operating Procedure Background Virtually all embedding media are water based. When a frozen tissue specimen embedded in such media is stored, over time the tissue and the medium will dehydrate and degrade. The Oil Bath Accessory Kit enables the user to coat the block face with the protective Oil that is maintained at -8°C in a bath inside the cryostat chamber.

  • Page 32: Use

    A tissue block can be stored in a freezer at -70°C for up to 1 year once thoroughly coated with the protective Oil. THE BLOCK MUST NOT BE PERMITTED TO THAW AND REFREEZE DURING STORAGE. THIS WILL DEGRADE THE QUALITY OF THE TISSUE IN THE BLOCK Storing a Tissue Block To store (short term in the cryostat or long term in a freezer) a frozen tissue block follow...

  • Page 33: Oil Bath Accessory Kit Troubleshooting Guide

    Oil Bath Accessory Kit Troubleshooting Guide ♦ The Oil inside the Oil Bath is frozen. Cause Solution Make sure the ECU is properly connected to an electrical source. Make sure the The ECU is not turned on. On/Off Switch on the ECU is in the On position (pushed in).
  • Page 34 ♦ The Oil Bath LED always displays a flashing red light. Cause Solution The temperature of the Oil inside the Oil Lower the cryostat temperature. bath is too warm. Call Instrumedics’ 800 customer service The ECU is malfunctioning. number and arrange for servicing of the ECU (1-800-237-2772).

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