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Processing

Embedding

QuestionAnswer
Supporting/embedding medium __ Holds cells & intercellular structures in proper relationship
Most popular embedding medium? Paraffin wax
As paraffin melting point increases ___ Paraffin becomes harder, provides better support
As paraffin melting point decreases __ Paraffin becomes softer, provides less support
Harder paraffin easier__ Thin sections
Softer paraffin easier __ Ribboning
Most common paraffin melting point? 55 °C to 58 °C
Sections may be processed at edge, soft in middle because? Sections too thick for good infiltration
Fix inadequately processed blocks b/c thick tissue? Trim unprocessed tissue off, embed, Trim sections down, xylene, tetrahydrofuran, xylene, re-infiltrate, embed
Carbowax embedding media are? Water-soluble waxes, polyethylene glycol+wax, blocks aren't permanent, dry up
Use of carbowax media doesn't require? Dehydration & clearing - waxes infiltrate directly from aq. fixatives, fat preserved
Carbowax won't infiltrate? Tissue w/ lots of fat, CNS infiltration v slow
Biggest problem w/ Carbowax + fix? Sections dissolve & "float out" on bath; To help add to bath : potassium dichromate+gelatin+water, diethylene glycol+formaldehyde +Carbowax+water
Carbowax sections are cut__? Chilled, 3 to 5 μm
Celloidin is? Any nitrocellulose embedding medium, use w/ any fixative
Most commonly used celloidin? Parlodion
Celloidin sections can be cut? Wet; keep wet w/ 80% until stained Dry: infiltrate w/ cedarwood oil first
Celloidin great for? CNS
Celloidin not often used b/c? Processing slow (weeks to months), uses very hazardous & explosive reagents, blocks can't be stored long
Celloidin processing does not require? Heat; less damage to tissue
GMA embedding medium Glycol methacrylate, acrylic resin, tolerates some water
GMA good for? Undecalcified bone, bone marrow, kidney, lymph node biopsies
GMA staining is__ More difficult than paraffin, some stains impossible b/c medium not removed from sections
Epoxy resins require__ Dehydrant & transitional fluid (like clearing agent for paraffin)
Most common epoxy transitional fluid? Propylene oxide
Most common epoxy resins? Araldite, Epon, & Spurr's, may cause dermatitis
Epoxy section thickness? Normal: 60-90 nm, thick: 0.5 μm
Agar & gelatin media used to? Make block w/ friable tissue Maintain specimen orientation during processing (double impregnation) Make frozen blocks (harden w/ formalin)
30% Sucrose use Infiltrate formalin-fixed, unprocessed tissue for frozen blocks
Tissue to be embedded diagonally? Hard tissues, e.g. bone
Tissue to be embedded on-edge? Tissues w/ wall or epithelium e.g. cysts, gallbladder, GI, skin, orient so knife through softest surface first
Tissues to be embedded in cross-section? Tubular structures
Forceps metastasis Fragments of tissue from one block are transferred to another on forceps, can lead to misdiagnosis, clean forceps between blocks
Blocks should be cooled quickly so__? Smaller paraffin crystals form, better support
Too much heat during processing or embedding may? Overfix/overharden tissue, negatively affect stainability
Celloidin processing procedure Dehydrate w/ equal parts 95% & 100%, Treat w/ equal mix 100%+ether, Infiltration w/ celloidin mixed w/ equal parts 100% & ether
Celloidin infiltration procedure Starts at 2% celloidin, gradually up to 12% to 14% celloidin Blocks hardened w/ chloroform
Microchatter Parallel tears in tissue potentially caused by overprocessed/overdehydrated tissue
Sponge artifact Caused by dry sponges being compressed into tissue
Best embedding medium for high-resolution light microscopy? GMA
Temperature of melted paraffin used for embedding? 2°C to 4°C above melting point
Polyethylene glycol can be used as embedding medium for? Lipids
OCT compound Optimal cutting temperature compound; water-soluble embedding medium, freeze quickly w/ liquid nitrogen, frozens
Other terms for embedding? Casting, blocking
Created by: CCF