Random

Nikita Gill (via untamedunwanted)

Part of my histology series

Aims of fixation

Prevention of bacterial growth and autolysis (cells being destroyed by their own enzymes) 

Prevent shrinkage, expansion i.e. change of shape during the subsequent processing and various tests

Allow staining of tissues to visualise various tissue components, cells, antigens etc. 

Must penetrate evenly and rapidly, and harden tissues

Proteins

 Fixatives have the properties of forming cross links between proteins to form a gel, ideally keeping everything as in vivo 

Soluble proteins are fixed to structural proteins & rendered insoluble, & whole structure given mechanical strength 

Classifications of fixatives

Aldehydes e.g. formaldehyde, glutaraldehyde (all non-coagulant) 

Oxidising agents: osmium tetroxide, potassium dichromate (non –coagulant) 

Protein-denaturing (coagulant): acetone, acetic acid, methyl alcohol, ethyl alcohol 

Physical: Heat, microwaves 

Miscellaneous: Mercuric chloride, picric acid, (coagulants) 

Aldehydes

FORMALDEHYDE 

(HCHO) 

Colourless gas, water soluble

In the lab: 10% formal saline/neutral buffered formalin

Acidic groups of the protein are unaffected by formaldehyde, basic groups are blocked. 

Does not precipitate DNA in solution. 

Most lipids are well preserved, though not necessarily fixed. 

Some phospholipids are very slowly dissolved by formalin. 

Formaldehyde useful for study of glycogen. 

GLUTARALDEHYDE 

for use with transmission Electron microscopy (TEM). 

Irreversible 

Fast fixation and penetration of tissue

Primary fixation with glutaraldeyhde; secondary fixation in osmium tetroxide to retain lipids 

Oxidising agents

OSMIUM TETROXIDE (OSO4 )

Occurs as pale yellow crystals 

Lipids; useful for TEM 

POTASSIUM DICHROMATE (K2 CR2 07 ) 

Soluble at around 10% in water at room temperature, but insoluble in absolute alcohol. 

Proteins converted to gel rather than precipitated. 

DNA dissolved; little effect on carbohydrates; useful for lipid demonstration. 

Has in the past been used for fixing tumours suspected of being pheochromocytomas. 

Protein Denaturing Reagents 

ETHYL ALCOHOL, METHYL ALCOHOL, ACETONE 

Mainly used for the fixation of cytological material. In some under fixed tissue specimens initially fixed with formalin may undergo some fixation by alcohol during the subsequent ‘paraffin processing’ 

precipitation, dehydration 

Lipids: not preserved/dissolved 

Carbohydrates: associated with proteins will also be precipitated 

Shrinkage, hardening, morphological considerations 

Temperature

TEMPERATURE FIXATION/STABILISATION 

Tissue placed in buffer/saline/isotonic solution 

useful for the rapid fixation of routine surgical specimens. 

For EM and some histochemistry best done at 0- 4C 

Freezing covered in tissue stabilisation post. 

Miscellaneous 

MERCURIC CHLORIDE (HG CL2 ) 

Secondary Fixative reacts and cross links proteins via the sulphydryl (-SH) group of cysteine amino acids 

Reputed to give superior morphology for H&E and special stains such as trichrome methods. 

feel free to change the pronouns and things to fit your muse!

“did you let _____ put worms in the blender again?”

“look alive, make yourself useful.”

“i’m sorry, why are you talking?”

“she shoots, she scores, unlike you, soccer boy.”

“you should ask him to the dance.”

“the girl is hot but she’s mad drama.”

“if your butt is suffering, then you are suffering.”

“that’s because you don’t have a soul.”

“sorry, you should take my jacket.”

“i will die before i go back to my daddy’s farm.”

“it’s obvious that your dad doesn’t really get you.”

“you’d make me proud if i actually cared about you.”

“you didn’t do anything. my dad married you.”

“i’m a star, a bright, gassy star.”

“_____, baby, your life sucks.”

“i was naked, of course he was flirting.”

“i’ll give _____ all the mothering you don’t have time for.”

“i’m not gonna let you not go to the dance without me. besides, we have six hours of shirtless team jacob so let’s just get this party started!”

“the chainsaw’s the best part!”

“i know your secret.”

“i will make your night. i promise.”

“don’t talk about her like that.”

“i’ll do whatever you want, okay? just leave ____ out of this.”

“i didn’t think i’d see you again.”

“they say that love tightens your pores.”

“you are so in over your head and you don’t even know it.”

“you always seem to disappear. i wish you didn’t do that.”

“don’t break my heart before i give it to you.”

“she’s not getting away with ruining your life.”

“you’re annoying and ugly but i sort of love you.”

“i’m nervous about tonight.”

“listen, he’s not falling for you, he’s falling for me.”

“you’re my best friend, i’d do anything for you.”