HT Unit 3: Fixation

define fixation

stabilization of tissue proteins to prevent autolysis and putrefaction

define autolysis

self digestion by endogenous enzymes

define putrefaction

bacterial decomposition

main purpose of fixation

preserve morphology by preventing autolysis and putrefaction

additive fixative do what

chemically bind to tissue proteins by altering the electrical charge which stabilizes the protein

non-additive fixative do what?

remove water which in turn shrinks and hardens tissue

coagulant fixative do what?

creates a mesh network which allows for faster penetration time

non-coagulant fixatives do what?

form gel which makes penetration to be slow

Example of non-additive fixatives?

Alcohols, Acetone, and Acetic Acid

Examples of additive fixatives?

(Everything else) Formaldehyde, Glutaraldehyde, Glyoxal, Mercury Chloride, Zinc Salts, Chromic acid, Picric acid, Osmium Tetroxide

Examples of coagulant fixatives?

Mercury Chloride, Acetic Acid, Chromic Acid, Zinc Salts, Alcohols, Picric Acid

examples of nom-coagulant fixatives?

formaldehyde, glutaraldehyde, glyoxal, osmium tetroxide

factors affecting fixation

temperature, size, fixative volume, time, penetration, pH, and osmolality

ideal tissue thickness?

less than 3mm

ideal fixative volume ration?

15-20x tissue volume

minimum fixation time in NBF?

6-8 hours

temperature for EM fixation?

0-4C

penetration fastes?

formaldehyde

penetration slowest?

picric acid

formaldehyde is additive or non-additive + coagulant, or non-coagulant?

additive, non-coagulant fo

formaldehyde stock concentration?

37-40%%

10% formalin contains ___% formaldehyde

4

formaldehyde forms ___ bridges to bind to proteins

methylene

what is added to formaldehyde to avoid polymerization?

methanol

unbuffered formalin forms what

formic acid

formalin pigment = aka

acid hematin

to remove formalin pigment you use what

alcoholic picric acid or alkaline alcohol

formaldehyde OSHA PEL, STEL, and action level

PEL = 0.75 ppm
STEL = 2.0 ppm
Action level = 0.5 ppm

glutaraldehyde is a

dialdehyde

glutaraldehyde is ____,_____

additive, non-coagulant

glutaraldehyde is used in

EM

T/F: glutaraldehyde gives false-positive PAS

True

glyoxal is a

dialdehyde

glyoxal dissolves ___, why?

eosinophils; due to its acidity which leaches iron

glyoxal does what to RBCs

lyses

glyoxal ineffective for ___

H.pylori

acetic acid is a ____,_____

coagulant, non-additive

acetic acid preserves ____

nucleoproteins

acetic acid does what to RBCs

swelling AND lyses

acetic acid is great to remove what?

microcalcifications

T/F: mercuric chloride is a strong coagulant which makes it a strong Stainer enhancer

True

Mercuric chloride does what to staining?

enhances

mercury pigment is removed with

iodine and thiosulfate

whats a replacement for mercury?

zinc

zinc salt is a _____,_____

coagulant, additive

con of zinc chloride?

causes precipitates which clogs processor

osmium tetroxide does what to lipids?

fixes them black

osmium tetroxide is used for

EM post-fixation

potassium dichromate <pH 3.5 =

coagulant

potassium dichromate > pH 3.5 =

non-coagulant

potassium dichromate has what type of pigment?

chrome pigment (yellow-brown)

chrome pigment is removed with

1% HCl in 70% alcohol

picric acid is ___ when dry

explosive

picric acid is both a fixative AND a ___

stain

T/F: You MUST wash out picric acid

True

B-5 contains

mercuric chloride, sodium acetate, formaldehyde

B-5 is excellent for

nuclear detail

Zenker does what to RBCs

lyses

Zenker is made up of what?

Mercuric chloride

Helly does what to RBCs

preserves

Bouin’s is a

trichrome

Bouin’s does what to RBCs

lyses

Gendre does what to glycogen

preserves

Hollande does what to RBCs

stabilizes

Zamboni is used for

EM

Orth is used for

chromaffin granules

Zinc formalin is used for

preservation of antigenicity

non-aqueous fixatives