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HISTOLOGICAL FIXATIVES

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ConferschemicalstABIlityonthetissue
Hardensthetissue(helpsfurtherhandling)
Haltsenzymeautolysis
Haltsbacterialputrefaction
Mayenhancelaterstainingtechniques
Introducesa"consistentartefact"

Classification:

Fixativesmaybeclassedasprecipitant(P)ornon-precipitant(NP)accordingtotheireffectontissueprotein.
Primaryfixativesare:
Aceticacid(NP)
Chromiumtrioxide(P)
Ethanol(P)
Formalin(NP)
Mercuricchloride(P)
Methanol(P)
Osmiumtetroxide(NP)
Picricacid(P)
Potassiumdichromate(NP)

Aceticalcohol-fixationtime1minute.

(Forsmears,cytospinpreparationsorfrozensections).

95%methanol-100ml

Glacialaceticacid-3ml.

Thesectionsshouldbewashedinwaterbeforestaining.

Bouin"sfluid-fixationtime6hours.

Saturatedaqueoussolutionofpicricacid-75ml

Formalin-25ml

Glacialaceticacid-5ml

Fixedtissueshouldbetransferredto70%alcohol.

Carnoy"sfluid-fixationtime1-3hours.

Ethanol-60ml

Chloroform-30ml

Glacialaceticacid-10ml

Fixedtissueshouldbeprocessedimmediatelyortransferredto80%alcohol.

Formolsublimate-fixationtime4-6hours.

Formalin-100ml

Mercuricchloride(saturatedaqueous)-900ml

Fixedtissueshouldbetransferredto80%alcohol.

Helly"sfluid-fixationtime12-24hours.

Stocksolution:-

Potassiumdichromate-25g

Mercuricchloride-50g

Sodiumsulphate-10g

Distilledwater-1000ml

Foruse:-

Stocksolution-100ml

Formalin-5ml

Thefixativesolutionshouldbemadeupjustbeforeuse.Fixedtissuemustbewashedfor24hoursinrunningtapwaterpriortoprocessing.

Neutralbufferedformalin-fixationtime12-24hours.

Formalin(40%aqueoussolutionofformaldehyde)-100ml

Sodiumdihydrogenorthophosphate(monohydrate)-4g

Disodiumhydrogenorthophosphate(anhydrous)-6.5g

Distilledwater-900ml

Thisfixativeissuitableformosthistologicalpurposes.Itistobepreferredtoformol-saline(asingle10%solutionofformalinin9%aqueousNaCl)asformalinpigmentisavoided.Specimensmaybestoredinthisfluid.Thesolutionisisotonic.

Michel"sfixativeforimmunoflourescence-fixationtime24-48hours.

Buffer:

0.81gpotassiumcitrate

0.0625gN-ethylmaleimide-HANDLEWITHCARE!

0.123gmagnesiumsulphate

100mlsdistilledwater

Beforeuseadd55gammoniumsulphateandallowtodissolve.

AdjustpHto7.0-7.2with1MKOH.

Placetissuebiopsiesinfixativefor24-48hours.Washtissuesinbuffer,threetimesover10minutes,andfreezeat-70^oC.

Paraformaldehyde-fixationtimedependsontechniquetofollow.

Sodiumdihydrogenorthophosphate2.26%-41.5ml

Sodiumhydroxide2.52%-8.5ml

Heatto60-80^oCinacoveredcontainer.

Add2g`Analar"paraformaldehydeandstiruntildissolved.Filter.

Zenker"sfluid-fixationtime4-24hours.

Distilledwater-950ml

Potassiumdichromate-25g

Mercuricchloride-50g

Glacialaceticacid-50g

Fixedtissueshouldbewashedovernightinrunningtapwaterbeforeprocessing.

REMOVALOFFIXATIONPIGMENTS

Formalinpigment

1.Dewaxthesections,rinsein100%alcohol,rinsein70%alcohol,rinseindistilledwater.

2.Treatinsaturatedalcoholicpicricacidfor30minutesto2hours.

3.Washwellinrunningtapwater.

4.Ifyellowstainingofthesectionpersistsrinseindilutelithiumcarbonate.

5.Rinseintapwater.

6.Continuewithmethod.

Mercurypigment

1.Dewaxthesections,rinsein100%alcohol,rinsein70%alcohol,rinseindistilledwater.

2.TreatinLugol"siodinefor2minutes.

3.Decolourisein5%sodiumthiosulphatefor5minutes.

4.Washwellinrunningtapwater.

5.Continuewithmethod.

Dichromatepigment

1.Dewaxthesections,rinsein100%alcohol,rinsein70%alcohol,rinseindistilledwater.

2.Treatin2%HClin70%alcohol16-24hours.

3.Rinseintapwater.

4.Continuewithmethod.

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HISTOLOGICAL&nbsp;FIXATIVES

HISTOLOGICAL FIXATIVES