Learn More About the Division
Contact Us
-
How to Submit Formalin Fixed Tissues
Fixation is the single most important preparative histological technique. The purpose of fixation is to preserve tissues permanently in as life-like a state as possible. Fixation should be carried out as soon as possible after removal of the tissues. Poor fixation cannot be remedied at any later stage and is the source of most of the problems we see in our lab. In our experience, under-fixation is a much greater problem than over-fixation. If the tissue is under-fixed, exposure to alcohols can dramatically alter antigen structure, modify morphology, and result in the loss of up to 40% of the protein content of the tissue. These effects are permanent and irretrievable
Supplies
- Tissue cassettes.
- Closed container with adequate fixative.
- Number two pencil or Histology pens.
- Properly trimmed tissues.
- Pathology Research Resources Histology Laboratory Submission form.
Procedure
1. Label side and top of cassette with a #2 pencil or histology pen (allow drying before place cassette in formalin). Please write legibly.
2. Tissues submitted should not be more than 3 mm thick. Take as reference a nickel for thickness and a post stamp for size.
3. Handle all tissues gently; grasp samples at the edges. Place tissues in labeled cassettes and be certain that the cassettes are securely closed.
4. Do not place more than three tissue samples per cassette.
5. Tiny pieces must be wrapped in lens papers or placed between sponges.
6. Place cassettes in fixative as soon as possible, Ensure container is properly closed and labeled containing: Name of fixative, time of collection and number of cassettes. If tissues are submitted without cassettes, do not place them in conical tubes or narrow mouth containers.
7. Tissue cassettes should be fully submerged in fixative to prevent partial fixation which will affect the quality of the sections. The ratio of fixative to tissue (volume: tissue) should be 10:1.
8. Complete the Pathology Research Resources submission form.
9. Discuss your needs with the histotechnologist and/or provide a written protocols specifying
- Type of tissue submitted
- Orientation of tissue
- Thickness of the sections
- Number of sections per slide
- Number of slides per block
- Amount of levels or series
- Special instructions
Notes
- Specimens smaller than 1mm. may not survive processing due to cell shrinking and the agitation and vacuum pressure involved during processing.
- Any tissues that contains bone or calcified tissue require special handling.
Questions? Please visit our lab located at RSMB 7th floor, room 7128 or call us at 305-243-3824
-
How to Submit Frozen Tissues
One of the benefits of frozen tissue samples is the time saved by omitting the initial fixation step required for paraffin-embedded tissue, which avoids the possible requirement to retrieve epitopes masked by formaldehyde cross-linking. Frozen tissue is cut on a cryostat and sections can be stored at -80 °C for up to 1 year.
Supplies
- Disposable embedding mold
- OCT medium
- Aluminum foil
- Freezing spray Liquid Nitrogen, or Dry Ice
- Forceps
- Petri Dish
- Kim wipes
- PBS
Procedure
1. Before you dissect the animal get one special container filled with liquid nitrogen and another container filled with some dry ice.
2. Label base mold and partially fill the mold with OCT.
3. Remove desired tissue and place in cold PBS and wash out the blood.
4. Transfer tissue to a clean Petri dish and use the Kim wipes to absorb the PBS on the tissue surface.
5. Place tissue in pre-labeled base molds filled with OCT. Try to arrange tissue flat in OCT near the bottom so tissue is easily exposed when sections are cut.
6. Use forceps to hold base mold edge and place the base mold into the surface of the liquid nitrogen and just let bottom of base mould to touch the surface of the nitrogen. Hold until the tissue solidifies (around 30 seconds). Note: If block is left in too long, it may crack.
7. Remove tissue from liquid nitrogen and place blocked tissue on dry ice. (Tissue may be kept in plastic container, plastic bags or wrapped in aluminum foil.)
8. Store frozen tissue block in -80°C freezer until sectioning.
9. Complete Pathology Research Resources submission form.
10. Transport samples to Pathology Research Resources Laboratory on dry ice.
11. Discuss your needs with the histotechnologist and/or provide a written protocol, providing:
- Thickness of sections
- Number of sections per slides
- Number of slides per sample
- Fixed or unfixed sections
- Stained or unstained sections
- Special instructions
Notes
The Investigator should contact the Lab to schedule a Frozen Section Service. At that time, arrangements can be made to immediately deliver the specimens for processing, if feasible. Alternatively, the specimens can be obtained and frozen for later processing.
Questions? Please visit our lab located at RSMB 7th floor, room 7128 or call us at 305-243-3824