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Staining Criteria Handbook

General Pathology (Routine Histopathology)

Neuropathology

Edition 4

November 2015

NEQMANMA003 General Pathology and Neuropathology Staining Criteria Handbook

Edition 004

2

Index

Page

Haematoxylin and Eosin Assessment Criteria 3

Special Stains A & B Assessment Criteria 6

List of Assessment Criteria 26

• Haematoxylin and Eosin

• Special Stains

Assessment Criteria Definitions 30


• Special Stains

Appendix 40

• Haematoxylin and Eosin Model Description

• Scoring System

• Scoring based on criteria

• UK NEQAS CPT Stain Repertoire


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Haematoxylin and Eosin

Assessment Criteria


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Haematoxylin and Eosin Assessment Criteria

Pre Microtomy Insufficient cellular features for assessment

Crush artefacts

Foam inset artefact

Red blood cell lysis

Poor chromatin detail

Cracking

Nuclear bubbling

Nuclear meltdown

Incorrect orientation suspected

Incorrect trimming suspected

Microtomy Chatter / vibration

Displacement

Folds / creases

Knife back debris

Knife marks

Lifting

Position on slide

Section too thick

Section too thin

Section thickness variable

Squames / floaters / fibres

Trimming artefact

Water bath bubbles

Staining Haematoxylin Intensity too strong

Haematoxylin Intensity too weak

Haematoxylin colour not blue

Haematoxylin background staining

Eosin Intensity too strong

Eosin Intensity too weak

Eosin colour

Eosin not selective

Uneven staining

Stain deposit present

Post Staining Air bubbles

Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax

Section wiped / section off slide

Tissue exposed

Water present

Contaminant on slide


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Description of Staining Results

Nuclei must be stained purple blue with haematoxylin. The intensity must be strong

enough allow clear demonstration of nuclear detail at a medium power, but not too

strong to cause a loss of the chromatin granularity or excessive cytoplasmic or

connective tissue staining.

Where the haematoxylin has been differentiated out, minimal cytoplasmic or

connective tissue background staining with haematoxylin must remain. This

background if present must not reduce the effectiveness of the nuclear demonstration

or affect the colour and selectiveness of the eosin.

The eosin should be selective enough to demonstrate different cellular components

such as collagen, cytoplasm, red blood cells, cellular granules, amyloid etc.

The intensity must be appropriate to the section thickness and the haematoxylin

intensity. Where the eosin is too weak it will fail to allow selective demonstration of

different components at low power. If the eosin intensity is too strong the colour and

detail of the nuclear stain will be obscured and selectivity will be reduced.

For extended description including pre Microtomy, Microtomy and post staining

please see model description in appendix.


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Special Stains A & B

Assessment Criteria


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Alcian Blue/PAS Assessment Criteria

Primary Stain Alcian Blue Intensity too strong

Alcian Blue Intensity too weak

Alcian Blue Colour

PAS Intensity too strong

PAS Intensity too weak

PAS Coloration

Not selective

Uneven Staining

Background

Deposit / precipitate present

Counterstain Nuclear Stain Intensity too strong

Nuclear Stain Intensity too weak

Nuclear Stain colour

Not selective

Uneven Staining


Post staining Air bubbles

Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Acid mucopolysaccharides should be coloured bright blue. Neutral mucopolysaccharides

should be bright magenta. Mixed mucins should be purple.

There should be clear distinction between acid, neutral and mixed mucins. Background

staining should be negligible.

A counterstain (typically haematoxylin) is considered optional. If present, it should be light

and even and should not mask or alter the primary stains. It should assist location.

Section quality and presentation should not impair the result.


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Amyloid (method for) Assessment Criteria

Primary Stain - Congo red (stain) colour

Bright field Intensity too strong

Intensity too weak

Not selective

Uneven Staining

Background


Primary Stain - No green birefringence on cross polarisation

Cross polarised light Birefringence intensity too weak

Birefringence intensity too strong

Birefringence not selective

Counterstain Nuclear stain intensity too strong

Nuclear stain intensity too weak

Nuclear stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Amyloid: There is no amyloid specific dye. However, when performed correctly the alkaline alcoholic

Congo red method is highly selective for binding to amyloid, when sections are prepared at the

appropriate thickness (~5 10 µm) and staining is performed stringently. Use of other dyes may produce

sub optimal or erroneous results.

Brightfield: Amyloid should stain pink to red (congophilic). Stain intensity is seldom homogeneous and

may differ in different tissues reflecting the deposition, abundance and underlying organisation of the

amyloid fibrils. Collagen and elastin may be weakly coloured. Background staining must be negligible.

Haematoxylin counterstain should be light, even and not mask or alter the primary stain; it should assist

location.

Cross Polarised Light: Amyloid stained pink / red in brightfield microscopy must also exhibit a bright

green colour in cross polarised light (green birefringence). Denser deposits of amyloid may exhibit

yellow green or bright yellow birefringence. The colour and/or intensity of the primary stain must not

mask the green birefringence of amyloid in cross polarised light. Collagen and elastin usually give pale

yellow to vivid white polarisation colours.


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Axonal Swelling (method for) Assessment Criteria

Primary Stain Intensity too strong

Intensity too weak

Stain colour

Axonal Swelling demonstration good

Axonal Swelling demonstration poor

Low power visibility

Not selective

Uneven Staining

Background


Counterstain Nuclear Intensity too strong

Nuclear Intensity too weak


Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Axonal swelling may be demonstrated by silver impregnation methods such as

Bielschowsky or using antibodies such as amyloid precursor protein (APP).

Swellings should be clearly stained, visible at low power, and discernible from other

structures.

Immunohistochemical methods require a light nuclear counterstain.



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Copper Associated Protein (CAP) Assessment Criteria


Intensity too weak

Stain colour

Not selective

Uneven Staining

Background


Counterstain Intensity too strong

Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



All copper associated protein should be stained and clearly identifiable.

HepBSA and elastin, if stained, should be readily distinguishable from CAP. Background

should be negligible.

If a counterstain is used it should not mask or modify the primary stain. It should assist

location.



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Diastase/PAS Assessment Criteria


Intensity too weak

Stain colour

Not selective

Uneven Staining

Background


Residual Glycogen




Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



The Schiff reaction is a histochemical method for aldehyde groups. Periodic acid oxidation

creates these groups on a variety of tissue structures notably glycogen, mucins, basement

membrane and fungi.

This modification uses the enzyme diastase (or amylase) to digest out any glycogen and is

normally used in conjunction with a straight PAS (not required for EQA unless requested).

Complete removal of glycogen and precise, complete demonstration of intestinal

mucopolysaccharide.

Membrane staining should not be confused with background, which should be minimal.

Counterstain (typically haematoxylin) should provide good colour contrast and assist

location.

Section quality and presentation must not impair the result.


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Elastin van Gieson Assessment Criteria


Intensity too weak

Stain Colour

No contrast

Fine Fibres

Not selective

Background

Uneven Staining


Counterstain Collagen Stain Intensity too strong

Collagen Stain Intensity too weak

Collagen Stain colour

Cytoplasm Stain Intensity too strong

Cytoplasm Stain Intensity too weak

Cytoplasm Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Elastin stains fall into two main groups, Haematoxylin based (e.g. Verhoeff) and

hydrophobic dye mixes (e.g. Miller’s and Weigert’s)

Precise staining of all coarse and fine elastin fibres.

The counterstain is a simple trichrome and there should be good colour separation

between muscle (yellow) and collagen (red).

Picro Sirius red is a suitable alternative, popular in the USA. The counterstain should

provide good colour contrast and assist location.



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Glial Fibres (method for) Assessment Criteria


Intensity too weak

Stain colour

Glial Fibre demonstration good

Glial Fibre demonstration poor


Not selective

Uneven Staining

Background



Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Glial fibres may be demonstrated using tinctorial methods such as PTAH or using antibodies

such as GFAP.

Astrocytic fibres should be clearly stained against a pale or clear background.

With PTAH, cell nuclei and myelin will also stain this requiring no further counterstaining.




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Gram Stain Assessment Criteria

Primary Stain Gram Positive Intensity too strong

Gram Positive Intensity too weak

Gram Positive Colour

Not selective

Uneven Staining

Background



Counterstain Gram Negative Intensity too strong

Gram Negative Intensity too weak

Gram Negative Colour

Not selective

Uneven Staining




Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Crystal Violet staining should render all Gram positive organisms blue/black without over or

under differentiation. There should be no violet background.

The “counterstain” serves to demonstrate all Gram negative organisms in a contrasting

colour and to assist location.

Most variations are based on the counterstain used and vary widely.



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Grocott Assessment Criteria

Primary Stain Organism Demonstration too strong

Organism Demonstration too weak

Organism Colour

Not selective

Uneven Staining

Background


Non Specific Silver Precipitate


Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



There should be dark grey/black staining of all fungal spores and/or hyphae or pneumocysts.

Where appropriate, internal structures of the organisms should be visible.

Impregnation of other structures, particularly collagen, should be negligible. There should be

no non specific silver precipitation.

The counterstain should provide good colour contrast and assist location, but must not mask

any delicately impregnated fungi.



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Haematoxylin / Van Gieson Assessment Criteria

Primary Stain Cytoplasm Stain Intensity too strong


Cytoplasm Stain colour

Collagen Stain Intensity too strong


Collagen Stain Colour

Not selective

Uneven Staining

Background




Nuclear Stain Colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



HVG is perhaps the simplest of trichrome methods which are designed to give colour

separation between smooth muscle and collagen.

The emphasis is on collagen, all of which should be stained bright red.

Muscle, erythrocytes and general cytoplasm (including hepatocytes) should be yellow.

Nuclear staining should be dark blue or black but must not influence the cytoplasmic

colouration by being under differentiated.



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Martius Scarlet Blue Assessment Criteria

Primary Stain Fibrin Demonstration too strong

Fibrin Demonstration too weak

Muscle Stain Intensity too strong

Muscle Stain Intensity too weak

Muscle Stain Colour




Not selective

Uneven Staining

Red blood cell colour

Background





Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



MSB is a specialised trichrome method designed to demonstrate fibrin of various ages in

colours from orange to red. Early fibrin can also stain yellow and old fibrin blue.

All fibrin should be brightly coloured and distinguishable from other structures.

As with other trichromes, there should be good colour separation between the dyes best

seen in red blood cells (yellow), muscle (red) and collagen (blue).

Nuclear staining should be dark blue or black.



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Masson Fontana (for melanin) Assessment Criteria

Primary Stain Melanin Demonstration too strong

Melanin Demonstration too weak

Not selective

Uneven Staining

Background



Incomplete Toning


Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



All melanin should be coloured black by silver impregnation.

The reaction product should be toned to distinguish it from unreacted melanin.

There should be no background staining arising from over impregnation and no non specific

silver precipitation.

The counterstain should provide good colour contrast and assist location.



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Myelin (method for) Assessment Criteria


Intensity too weak

Stain colour

Myelin demonstration good

Myelin demonstration poor


Not selective

Uneven Staining

Background



Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Axonal myelin sheaths may be demonstrated by tinctorial methods such as Luxol fast blue

(LFB) or using antibodies such as myelin basic protein.

All myelin should be clearly stained against a pale/clear background. Fine fibres should be

discernible.

LFB should be counterstained, traditionally with cresyl fast violet.



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Neurofibrillary Tangles Assessment Criteria


Intensity too weak

Stain colour

Tangle demonstration good

Tangle demonstration poor


Not selective

Uneven Staining

Background



Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Neurofibrillary Tangles may be demonstrated quite specifically using silver impregnation

methods such as Gallyas or using antibodies such as Tau or Ubiquitin

All tangles should be clearly stained, visible at low power, and discernible from other

structures.




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Nissl Substance Assessment Criteria


Intensity too weak

Stain colour

Nissl demonstration good

Nissl demonstration poor


Not selective

Uneven Staining

Background



Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



All nissl granules in neuronal cell bodies should be clearly stained purple / blue.

Cell nuclei should also be demonstrated. There should be no negligible background and no

stain precipitate.

This method may also be employed as a counterstain for other techniques.



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Perls’ Prussian Blue Assessment Criteria


Intensity too weak

Stain Colour

Not selective

Uneven Staining

Background


Unreacted iron


Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



A histochemical reaction for ferric iron adapted for the demonstration of haemosiderin

Prussian Blue colouration of haemosiderin should not be modified by counterstain or

underlying, unreacted pigment.

The counterstain should provide good colour contrast and assist location.



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Reticulin (silver method for) Assessment Criteria

Primary Stain Reticulin Demonstration too strong

Reticulin Demonstration too weak

Not selective

Uneven Staining

Background



Incomplete Toning


Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



This method is primarily a low power scanning method so contrast is all important. There

are many methods, the most popular in the UK being Gordon and Sweet. In Europe and the

US Gomori is more popular. The latter impregnates nuclei which, when done well, renders

counterstaining unnecessary.

The main variation in method is toning. Without it, collagen would remain golden brown

and cell cytoplasm a pale yellow. Again, counterstaining is unnecessary if the rest of the

method is done well.

Reticulin should be black, under impregnation will result in incomplete staining, over

impregnation yields knobbly fibres. Fine reticulin fibres should be clearly seen at high

power.

There should be no non specific silver deposits and staining of cytoplasmic structures is

undesirable.

Counterstaining, if used, should provide good contrast and assist location. Gomori and its

variants may render toned collagen rose-pink.



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Senile Plaques (method for) Assessment Criteria


Intensity too weak

Stain colour

Plaque demonstration good

Plaque demonstration poor


Not selective

Uneven Staining

Background



Incomplete Toning


Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Plaques may be demonstrated quite specifically using silver impregnation methods such as

Haga or by virtue of their amyloid component using antibodies such as β amyloid A4.

All plaques should be clearly stained against a pale or clear background and be visible at

low power. The dystrophic neuritis of neuritic plaques may also be demonstrated.

Silver methods generally create a yellow background which requires no further

counterstaining.

Immunohistochemical methods require a light nuclear counterstain



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Trichrome (not HVG) Assessment Criteria

Primary Stain Muscle Stain Intensity too strong


Muscle Stain Colour




Not selective

Uneven Staining


Background





Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



Trichrome methods are designed to give colour separation between smooth muscle and

collagen. There are a variety of methods; Masson’s being the most popular in the UK.

All muscle should be stained red, collagen blue or green and nuclei blue/black.

The collagen (milling) dye may be green or blue.

Nuclear staining should be dark blue/black.


NB: HVG is assessed separately and will not be accepted if submitted as a “standard”


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Ziehl – Neelsen Assessment Criteria

Primary Stain Bacilli stain Intensity too strong

Bacilli stain Intensity too weak

Bacilli Colour

Not selective

Uneven Staining

Background




Intensity too weak

Stain colour

Not selective

Uneven Staining



Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue Damage

Tissue exposed

Water present



A stain used for Mycobacterium tuberculosis and closely related organisms

All mycobacteria, including isolated bacilli, should be sharply stained magenta/red by carbol

fuchsin with minimal background staining.

The counterstain should be light, even and predominantly nuclear. It should provide good

colour contrast assist location.

There should be no dye precipitation and the tissue should not be damaged by heating.



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List of

Assessment Criteria


• Special Stains


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Haematoxylin and Eosin Assessment Criteria

Pre Microtomy Insufficient cellular features for assessment

Crush artefacts

Foam inset artefact

Red blood cell lysis

Poor chromatin detail

Cracking

Nuclear bubbling

Nuclear meltdown

Incorrect orientation suspected

Incorrect trimming suspected

Microtomy Chatter / vibration

Displacement

Folds / creases

Knife back debris

Knife marks

Lifting

Position on slide

Section too thick

Section too thin

Section thickness variable

Squames / floaters / fibres

Trimming artefact

Water bath bubbles

Staining Haematoxylin Intensity too strong

Haematoxylin Intensity too weak

Haematoxylin colour not purple blue

Haematoxylin background staining

Eosin Intensity too strong

Eosin Intensity too weak

Eosin colour

Eosin not selective

Uneven staining

Stain deposit present


Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue exposed

Water present



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Special Stains Assessment Criteria

Primary Stain Alcian Blue Colour

Alcian Blue Intensity too strong

Alcian Blue Intensity too weak

Amyloid Bright field Background

Amyloid Bright field Deposit / Precipitate present

Amyloid Bright field Intensity too strong

Amyloid Bright field Intensity too weak

Amyloid Bright field Stain colour

Axonal Swelling demonstration good

Axonal Swelling demonstration poor

Bacilli Colour

Bacilli stain Intensity too strong

Bacilli stain Intensity too weak

Background

Birefringence intensity too weak

Birefringence colour

Birefringence not selective




Cytoplasm Stain Colour




Fibrin Demonstration too strong

Fibrin Demonstration too weak

Fine Fibres

Glial Fibre demonstration poor

Glial Fibre demonstration good

Gram Positive Colour

Gram Positive Intensity too strong

Gram Positive Intensity too weak

Incomplete Toning

Intensity too strong

Intensity too weak


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Primary Stain continued Low power visibility

Melanin Demonstration too strong

Melanin Demonstration too weak

Muscle Stain Colour

Muscle Stain Intensity too strong


Myelin demonstration poor

Myelin demonstration good

Nissl demonstration good

Nissl demonstration poor

No green birefringence on cross polarisation

No Contrast


Not selective

Organism Colour

Organism Demonstration too strong

Organism Demonstration too weak

PAS Coloration

PAS Intensity too strong

PAS Intensity too weak

Plaque demonstration good

Plaque demonstration poor


Residual Glycogen

Reticulin Demonstration too strong

Reticulin Demonstration too weak

Stain colour

Tangle demonstration good

Tangle demonstration poor

Uneven Staining

Unreacted iron


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Counterstain Collagen Stain Colour



Cytoplasm Stain Colour




Gram Negative Colour

Gram Negative Intensity too strong

Gram Negative Intensity too weak

Intensity too strong

Intensity too weak


Not selective


Nuclear Stain Intensity too strong


Stain colour

Uneven Staining


Air drying artefact

Excessive mountant

Mountant shrinkage

Residual wax


Tissue exposed

Water present



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Assessment Criteria

Definitions


• Special Stains


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Haematoxylin and Eosin Criteria Definitions

Pre Microtomy The material supplied must be paraffin wax processed and have sufficient variety of cellular

features, in particular nuclei to allow assessment.

Insufficient cellular features for assessment - The material supplied does not have sufficient variety

of cellular features, in particular nuclei to allow assessment

Artefacts, such as crush effects and foam insert impressions should not be introduced during the

laboratory handling and preparation stages.

Crush artefacts - distortion of nuclei to give a thin elongated appearance because of crushing prior

to fixation

Foam inset artefact - triangular holes on the outer surface of biopsies produced by the surface of

foam cassette inserts

The tissue must show no evidence of being inadequately fixed or having had delayed fixation and

there must be a clear demonstration of the chromatin detail within the nuclei at medium and low

power and little cell lysis.

Red blood cell lysis -loss of structure and shape and fusion of red cells

Poor chromatin detail - insufficient chromatin clumping, granularity or perinuclear chromatin

There must be no evidence of poor paraffin processing. Epithelial cells groups and connective

tissue components must not appear to be separated by cracking. There should be no disruption of

the nuclear membrane resulting in pale, fused nuclear staining (nuclear meltdown). The

appearance of bubble like artefacts over the nuclei will be noted but no marks will be deducted.

Cracking - excessive separation of epithelial cells, lymphoid cells or connective tissue

Nuclear bubbling - the appearance of a vacuole or bubble shape over and within nuclei

Nuclear meltdown - loss of a distinct nuclear membranes and pale staining in nuclei

If the biopsy is suspected of being orientated in a way that fails to demonstrate the cell types or

layers appropriately this will be noted but no deduction will be made from the score. Similarly if it

is felt that the tissue has not been trimmed to full face, or has been trimmed past full face this will

be noted but no deduction will be made.

Incorrect orientation - suspected lack of all the cell layers expected in a tissue type

Incorrect trimming - suspected lack of areas of epithelium or other which could be expected


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Microtomy

The section must be of a thickness appropriate to the tissue type. The section is too thick if it is

difficult to focus on a single layer of nuclei, or too thin if staining intensity is compromised. It must

be free from variations in thickness, creases, folds, scores and contaminants. No accumulation or

fragments of tissue should be seen over or between sections. The section should be flat, with no

lifted areas. There should be no holes, tearing or damage either as a result of trimming or

sectioning or cover-slipping. The section must be positioned on the slide so that microscopy is not

compromised.

Chatter / vibration - very closely placed variation in thickness causing stripes of alternate staining

intensity at right angles to direction of sectioning

Displacement - cells that become detached and move over other areas of section

Folds / creases - creases and folds in the section giving doubles layers that lift from the slide and

stain intensely

Knife back debris - accumulation of cells that are smeared on the edge of the knife and deposited

between alternate sections

Knife marks - scores and scratches at parallel to the direction of sectioning

Lifting - areas of section that lift and do not lie flat

Position on slide - section positioned appropriately on slide for full visualisation

Section too thick - unable to focus on a single cell layer

Section too thin - loss of stain intensity and contrast because of thinness of section

Section thickness variable - varying thickness in different areas of the section, recognised by varying

dye intensity

Squames / floaters / fibres -contamination by material that is not in the block, usually above the

section

Trimming artefact - tears and rough edged holes, often with lifted edges, within the section

Water bath bubbles circular areas of lifting, often cracked and intensely stained


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Staining

Nuclei must be stained purple/ blue with haematoxylin. The intensity must be strong enough

allow clear demonstration of nuclear detail at a medium power, but not too strong to cause a loss

of the chromatin granularity or excessive cytoplasmic or connective tissue staining. Where the

haematoxylin has been differentiated out, minimal cytoplasmic or connective tissue background

staining with haematoxylin must remain. This background if present must not reduce the

effectiveness of the nuclear demonstration or affect the colour and selectiveness of the eosin.

Haematoxylin Intensity too strong - a loss of the chromatin granularity or excessive cytoplasmic or

connective tissue staining

Haematoxylin Intensity too weak - intensity must be strong enough to allow clear demonstration of

nuclear detail at a medium power

Haematoxylin colour not purple - blue Nuclei must be stained purple/ blue with haematoxylin

Haematoxylin background staining - where the haematoxylin has been differentiated out, minimal

cytoplasmic or connective tissue background staining with haematoxylin must remain. If present

must not reduce the effectiveness of the nuclear demonstration or affect the colour and

selectiveness of the eosin

The eosin should be selective enough to demonstrate different cellular components such as

collagen, cytoplasm, red blood cells, cellular granules, amyloid etc. The intensity must be

appropriate to the section thickness and the haematoxylin intensity. Where the eosin is too weak

it will fail to allow selective demonstration of different components at low power. If the eosin

intensity is too strong the colour and detail of the nuclear stain will be obscured and selectivity will

be reduced.

Eosin Intensity too strong - the intensity must be appropriate to the section thickness and the

haematoxylin intensity. If the eosin intensity is too strong the colour and detail of the nuclear stain

will be obscured and selectivity will be reduced

Eosin Intensity too weak - the intensity must be appropriate to the section thickness and the

haematoxylin intensity. Where the eosin is too weak it will fail to allow selective demonstration of

different components at low power

Eosin colour - the cytoplasm is not stained the appropriate colour based on the method employed

and the expected staining results . Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven

washing. Dehydrated incorrectly.

Eosin not selective - the eosin should be selective enough to demonstrate different cellular

components such as collagen, cytoplasm, red blood cells, cellular granules, amyloid etc

All staining should be even across the section and there should be no dye deposits or precipitate.

Uneven staining - all staining should be even across the section

Stain deposit present -dye deposits or precipitate


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Post Staining

Coverslipping must be free from air bubbles, air drying artefact or drying back. There should be no

excessive mountant, the sections must be totally covered and there must be no evidence of

incomplete dewaxing or dehydration. The difficulties of processing and sectioning bony tissue will

be taken into account when assessing.

Air bubbles - air bubbles within the mountant

Air drying artefact - retractile areas and tissue damage

Excessive mountant - mountant outside the coverslip

Mountant shrinkage - areas of mountant have dried back from the coverslip edges

Residual wax - retractile areas of undissolved wax are visible

Section wiped / section off slide - section scratched by hand or off edge of slide

Tissue exposed - the section is not covered by the coverslip

Water present - droplets of water under the coverslip

Contaminant on slide - Contamination by non biological or biological material, excluding dye /

reagent deposits, usually above the tissue section between the section and the coverslip. For

example, squames / floaters / fibres/ pencil or ink deposits


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Special Stains Criteria Definitions

Primary Stain

Alcian Blue Colour - the acid mucin is not bright blue, and the mixed mucins are not purple. Old

dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing. Dehydrated incorrectly

Alcian Blue Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with the

nuclear stain or staining areas that should be another colour. Other detail may be missing or

selectivity could be reduced.

Alcian Blue Intensity too weak - where the stain is too weak it will fail to allow the clear, selective

demonstration of different components at low power the intensity must be appropriate to the

section thickness.

Amyloid Bright field Background - Background staining present, reducing contrast of the nuclei or

obscuring detail or affecting colour of amyloid deposits. Caused by staining too long, not

differentiating enough or non-selective staining, or breakdown products in old stain.

Amyloid Bright field Deposit / Precipitate present - Random irregular or crystalline deposits above

and around amyloid deposits. Old or unfiltered dye.

Amyloid Bright field Intensity too strong - Stain intensity is so strong that it obscures / interferes

with the nuclear counterstain, or staining areas that should be another colour. Other detail may be

missing or selectivity could be reduced.

Amyloid Bright field Intensity too weak - Stain intensity is so weak that it is failing to demonstrate

amyloid deposits clearly.

Amyloid Bright field Stain colour – Amyloid is not stained the appropriate colour based on the

method employed and the expected staining results. Old dye. Mixed incorrectly. Not evenly

applied to slide. Uneven washing. Dehydrated incorrectly.

Axonal Swelling demonstration good – swellings should be clearly stained, visible and discernible

from other structures.

Axonal Swelling demonstration poor - ineffective demonstration prevents successful visualisation.

Bacilli Colour - the bacilli are not stained magenta / red. Old dyes. Mixed incorrectly. Not evenly

applied to slide. Uneven washing. Dehydrated incorrectly.

Bacilli stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with the

clear, selective demonstration of bacilli or staining areas that should be another colour. Other detail

may be missing or selectivity could be reduced.

Bacilli stain Intensity too weak - where the stain is too weak it will fail to allow the clear, selective


section thickness.


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Primary Stain continued

Background - background staining present, reducing contrast of the nuclei or obscuring detail or

affecting colour of another feature. Caused by staining too long, not differentiating enough or non-

selective breakdown products in old stains.

Birefringence intensity too weak – Birefringence intensity is so weak in cross polarised light that it is

failing to demonstrate amyloid deposits clearly.

Birefringence colour – Birefringence is not the appropriate colour based on the method employed

and the expected results in cross polarised light. Incorrect / non-stringent method. Old dye. Mixed

incorrectly. Not evenly applied to slide. Uneven washing. Dehydrated incorrectly.

Birefringence not selective – Birefringence not selective of different tissue components in cross

polarised light. Incorrect / non-stringent method. Old dye. Mixed incorrectly. Not evenly applied to

slide. Uneven washing. Dehydrated incorrectly.

Collagen Stain Colour - the collagen is not stained the appropriate colour based on the method

employed and the expected staining results . Old dyes. Mixed incorrectly. Not evenly applied to


Collagen Stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with

demonstration of collagen and different components or staining areas that should be another

colour. Other detail may be missing or selectivity could be reduced.

Collagen Stain Intensity too weak- where the stain is too weak it will fail to allow the clear, selective

demonstration of collagen and different components at low power the intensity must be

appropriate to the section thickness.

Cytoplasm Stain Colour - the cytoplasm is not stained the appropriate colour based on the method



Cytoplasm Stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering

with demonstration of different cytoplasmic components. Other detail may be missing or selectivity

could be reduced.

Cytoplasm Stain Intensity too weak- where the stain is too weak it will fail to allow the clear,

selective demonstration of different cytoplasmic components at low power. The intensity must be


Deposit / Precipitate present- random irregular or crystalline deposits above and around the cells.

Old or unfiltered dyes

Fibrin Demonstration too strong - stain intensity is so strong that it is obscuring/ interfering with

demonstration of fibrin and different components or staining areas that should be another colour.

Other detail may be missing or selectivity could be reduced.

Fibrin Demonstration too weak- where the stain is too weak it will fail to allow the clear, selective

demonstration of fibrin and different components at low power the intensity must be appropriate to

the section thickness.


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Fine Fibres – effective demonstration allows fine fibres to be visualised successfully.

Glial Fibre demonstration good – fibres should be clearly stained against a pale or clear background,

and visible and discernible from other structures.

Glial Fibre demonstration poor - ineffective demonstration prevents successful visualisation.

Gram Positive Colour - the gram positive organisms are not stained blue / black. Old dyes. Mixed

incorrectly. Not evenly applied to slide. Uneven washing. Dehydrated incorrectly

Gram Positive Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of bacilli or staining areas that should be another colour. Other

detail may be missing or selectivity could be reduced.

Gram Positive Intensity too weak- where the stain is too weak it will fail to allow the clear, selective

demonstration of bacilli at low power the intensity must be appropriate to the section thickness.

Incomplete Toning – colouration impairs clear visualisation / demonstration.

Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with the nuclear

stain or staining areas that should be another colour. Other detail may be missing or selectivity could

be reduced.

Intensity too weak - stain intensity is so weak that it is failing to demonstrate tissue components

clearly

Low power visibility - where it is effective and precise to allow the clear, selective demonstration of

different components at low power. The intensity must be appropriate to the section thickness.

Melanin Demonstration too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of melanin or staining areas that should be another colour. Other


Melanin Demonstration too weak- where the stain is too weak it will fail to allow the clear, selective

demonstration of melanin at low power the intensity must be appropriate to the section thickness.

Muscle Stain Colour - the muscle is not stained the appropriate colour based on the method



Muscle Stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of muscle or staining areas that should be another colour. Other


Muscle Stain Intensity too weak- where the stain is too weak it will fail to allow the clear, selective

demonstration of muscle at low power the intensity must be appropriate to the section thickness.


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Myelin Demonstration good – myelin should be clearly stained against a pale or clear background,

and discernible from other structures.

Myelin Demonstration poor - ineffective demonstration prevents successful visualisation.

Nissl demonstration good– myelin should be clearly stained against a pale or clear background, and

discernible from other structures.

Nissl demonstration poor - ineffective demonstration prevents successful visualisation.

No green birefringence on cross polarisation - Amyloid deposits that are stained pink/red in

brightfield microscopy must also exhibit green birefringence (strong) in cross polarised light

No contrast – colouration impairs clear visualisation / demonstration of target cells, tissue

component or organism.

Non Specific Silver Precipitate - deposit / precipitate present as deposits above and around the

cells. Old or unfiltered dyes.

Not selective - Not selective of different cell types or tissue components. Old dyes. Mixed


Organism Colour - the organisms according to the staining results of the method employed. Old

dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing. Dehydrated incorrectly

Organism Demonstration too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of organisms or staining areas that should be another colour.

Other detail may be missing or selectivity could be reduced.

Organism Demonstration too weak- where the stain is too weak it will fail to allow the clear,

selective demonstration of organisms at low power the intensity must be appropriate to the section

thickness.

PAS Coloration - the neutral mucins are not bright magenta, and the mixed mucins are not purple.

Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing. Dehydrated incorrectly

PAS Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with the nuclear

stain or staining areas that should be another colour. Other detail may be missing or selectivity could

be reduced.

PAS Intensity too weak- where the stain is too weak it will fail to allow the clear, selective


section thickness.

Plaque Demonstration good – plaques should be clearly stained against a pale or clear background,

and discernible from other structures.

Plaque Demonstration poor - ineffective demonstration prevents successful visualisation.


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Red blood cell colour – red blue cells must be stained according to the staining results of the

method employed.

Residual Glycogen – residual glycogen present in tissue components after digestion, where glycogen

should no longer be demonstrated.

Reticulin Demonstration too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of thick and / or fine reticulin fibres or staining areas that should

be another colour. Other detail may be missing or selectivity could be reduced.

Reticulin Demonstration too weak- where the stain is too weak it will fail to allow the clear,

selective demonstration of different thick and / or fine at low power the intensity must be


Stain Colour –is not stained the appropriate colour based on the method employed and the

expected staining results . Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing.

Dehydrated incorrectly.

Tangles Demonstration good – neurofibrillary tangles should be clearly stained against a pale or

clear background, and discernible from other structures.

Tangles Demonstration poor - ineffective demonstration prevents successful visualisation.

Uneven staining - varying intensity of staining in different parts of the slide preparation. Dyes

applied but not spread evenly. Uneven dehydration.

Unreacted iron– a significant amount of haemosiderin has not reacted with the Perls reagent and

remains brown in colour.

Counterstain

Collagen Stain Colour - the collagen is not stained the appropriate colour based on the method



Collagen Stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with

demonstration of collagen and different components or staining areas that should be another

colour. Other detail may be missing or selectivity could be reduced.

Collagen Stain Intensity too weak- if the stain is too weak it will fail to allow the clear, selective

demonstration of collagen and different components at low power the intensity must be



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Counterstain continued

Cytoplasm Stain Colour - the cytoplasm is not stained the appropriate colour based on the method



Cytoplasm Stain Intensity too strong - stain intensity is so strong that it is obscuring/ interfering

with demonstration of different cytoplasmic components. Other detail may be missing or selectivity

could be reduced.

Cytoplasm Stain Intensity too weak- if the stain is too weak it will fail to allow the clear, selective

demonstration of different cytoplasmic components at low power. The intensity must be


Deposit / Precipitate present - Deposit / precipitate present - random irregular or crystalline

deposits above and around the cells. Old or unfiltered dyes.

Gram Negative Colour - the gram negative organisms are not stained according to the colour of the

counterstain employed. Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing.

Dehydrated incorrectly

Gram Negative Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with

the clear, selective demonstration of bacilli or staining areas that should be another colour. Other


Gram Negative Intensity too weak- where the stain is too weak it will fail to allow the clear,

selective demonstration of bacilli at low power the intensity must be appropriate to the section

thickness.

Intensity too strong - stain intensity is so strong that it is obscuring/ interfering with the nuclear

stain or staining areas that should be another colour. Other detail may be missing

Intensity too weak - stain intensity is so weak that it is failing to demonstrate tissue components

clearly

Low power visibility - where it is effective and precise to allow the clear, selective demonstration of

different components at low power. The intensity must be appropriate to the section thickness.

Not selective - Not selective of different cell types or tissue components. Old dyes. Mixed


Nuclear Stain Colour – nuclei are not stained the appropriate colour based on the method employed

and the expected staining results . Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven

washing. Dehydrated incorrectly.

Nuclear Stain Intensity too strong - Obscuring chromatin detail or staining non-nuclear features.

Usually caused by under differentiation/ or staining for too long. If so there may also be background

staining of cytoplasm


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Counterstain continued

Nuclear Stain Intensity too weak - Failing to clearly show chromatin detail. Nuclei don’t stand out

well on low power and are feint relative to the background. Old dyes or too short staining time or

over differentiated

Stain Colour –is not stained the appropriate colour based on the method employed and the

expected staining results . Old dyes. Mixed incorrectly. Not evenly applied to slide. Uneven washing.

Dehydrated incorrectly.

Uneven staining - varying intensity of staining in different parts of the slide preparation. Dyes

applied but not spread evenly. Uneven dehydration.

Post Staining

Air bubbles - air bubbles within the mountant

Air drying artefact - retractile areas and tissue damage

Excessive mountant - mountant outside the coverslip

Mountant shrinkage - areas of mountant have dried back from the coverslip edges

Residual wax - retractile areas of un-dissolved wax are visible

Section wiped / section off slide - section scratched by hand or off edge of slide

Tissue exposed - the section is not covered by the coverslip

Water present - droplets of water under the coverslip

Contaminant on slide - Contamination by non biological or biological material, excluding dye /

reagent deposits, usually above the tissue section between the section and the coverslip. For

example, squames / floaters / fibres/ pencil or ink deposits


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Appendix 1

• Haematoxylin and Eosin Model Description

• Scoring guidelines

• Scoring based on criteria

• UK NEQAS CPT Stain Repertoire


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Haematoxylin and Eosin Model Description

The material supplied must be paraffin wax processed and have sufficient variety of cellular features, in

particular nuclei to allow assessment. Artefacts, such as crush effects and foam insert impressions should

not be introduced during the laboratory handling and preparation stages.

The tissue must show no evidence of being inadequately fixed or having had delayed fixation and there

must be a clear demonstration of the chromatin detail within the nuclei at medium and low power and

little red cell lysis.

There must be no evidence of poor paraffin processing. Epithelial cells groups and connective tissue

components must not appear to be separated by cracking. There should be no disruption of the nuclear

membrane resulting in pale, fused nuclear staining (nuclear meltdown). The appearance of bubble like

artefacts over the nuclei will be noted but no marks will be deducted.

If the biopsy is suspected of being orientated in a way that fails to demonstrate the cell types or layers

appropriately this will be noted but no deduction will be made from the score. Similarly if it is felt that the

tissue has not been trimmed to full face, or has been trimmed past full face this will be noted but no

deduction will be made.

The section must be of a thickness appropriate to the tissue type. The section is too thick if it is difficult to

focus on a single layer of nuclei, or too thin if staining intensity is compromised. It must be free from

variations in thickness, creases, folds, scores and contaminants. No accumulation or fragments of tissue

should be seen over or between sections. The section should be flat, with no lifted areas. There should

be no holes, tearing or damage either as a result of trimming or sectioning or coverslipping. The section

must be positioned on the slide so that microscopy is not compromised.

Nuclei must be stained purple blue with haematoxylin. The intensity must be strong enough allow clear

demonstration of nuclear detail at a medium power, but not too strong to cause a loss of the chromatin

granularity or excessive cytoplasmic or connective tissue staining. Where the haematoxylin has been

differentiated out, minimal cytoplasmic or connective tissue background staining with haematoxylin must

remain. This background if present must not reduce the effectiveness of the nuclear demonstration or

affect the colour and selectiveness of the eosin.

The eosin should be selective enough to demonstrate different cellular components such as collagen,

cytoplasm, red blood cells, cellular granules, amyloid etc. The intensity must be appropriate to the

section thickness and the haematoxylin intensity. Where the eosin is too weak it will fail to allow selective

demonstration of different components at low power. If the eosin intensity is too strong the colour and

detail of the nuclear stain will be obscured and selectivity will be reduced.

All staining should be even across the section and there should be no dye deposits or precipitate.

Coverslipping must be free from air bubbles, air drying artefact or drying back. There should be no

excessive mountant, the sections must be totally covered and there must be no evidence of incomplete

dewaxing or dehydration. The difficulties of processing and sectioning bony tissue will be taken into

account when assessing.


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Scoring Guidelines

Each pair of assessors complete an assessment form and data from these forms are converted into a

mark out of 5, from each assessor. The mark out of 5 from each assessor is based on the criteria for

a given method.

Guide lines for individual assessors mark (out of 5)

where 0 – non submission, 1- Fail, 2 - Borderline Fail, 3 - Pass, 4 – Good, 5 - Excellent

0 – non submission

1 – Fail - No staining demonstrated based on the method employed and the expected staining

results

2 – Borderline Fail - unsatisfactory demonstration based on the method employed, with

expected staining results being inappropriate

3 – Pass - appropriate demonstration based on the method employed and the expected

staining results, although improvements need to be made in the staining.

4 – Good – good appropriate demonstration based on the method employed and the

expected staining results

5 – Excellent – excellent demonstration based on the method employed and the expected

staining results

Each assessor submits their mark out of 5 based on the criteria for a given method, giving a

total score for the submitted slide out of 10.

Guide lines for total score (out of 10)

Score <5 - A score of less than 5 / 10 is given for poor staining, where the participant has failed

to clearly demonstrate the expected results.

Score 5/6 – a score of 5 or 6 / 10 is a pass. Whilst the staining appropriately demonstrates the

expected staining results, staining is suboptimal and improvements are still required overall.

Score 7/8 – a score of 7 or 8 / 10 shows good appropriate demonstration of the expected

results, and an acceptable level of quality.

Score 9/10 – a score of 9 or 10 / 10 shows excellent appropriate demonstration of the

expected results, and a high level of quality.

NB. Any slides which score a mark of 4 or below are passed to a secondary assessor for further assessment

before a final score is issued. If there is a discrepancy of 2 between the assessing pair e.g. 3 & 5, the slide

will be passed for secondary assessing. If there is a discrepancy of pass / fail between the assessing pair, the

slide will be passed for secondary assessing.


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Scoring based on criteria

Below is a generic ideal score for haematoxylin & eosin, and special stains.

It is intended purely as a guide for laboratories and assessors. Slides may not achieve all of the points

listed and may have elements which span several score boundaries.

Score 5 - Excellent Excellent nuclear and tissue constituent staining in a suitable preparation

allowing full visualisation of nuclear and component features within the

tissue.

• Nuclear Staining intensity which demonstrates the chromatin detail clearly.

• Staining colour, intensity and balance allows clear distinction between nuclear detail and

other non-nuclear features.

• Primary stain and counterstain demonstrates the appropriate tissue constituents depending

on the method being employed.

• The cytoplasm must show appropriate colour spectrum.

• Counterstain intensity stain intensity does not obscure / interfere with the nuclear stain or

staining areas that should be another colour. Intensity is such that it is allows clear

demonstration of tissue components.

• Demonstration is appropriate based on the method employed and the expected staining

results.

• Even staining across the tissue section, with minimal background staining and no dye

deposits or precipitate.

• Suitable preparation to allow clear observation of the appropriate tissue constituents

depending on the method being employed.

• There are no microtomy or processing irregularities.

• Dehydration, covering and labelling does not impair the visualisation of the tissue and its

components.

• The overall preparation and staining is excellent and allows full visualisation of the tissue and

its components.

Score 4 - Good Good nuclear and tissue constituent staining in a suitable preparation to

allow visualisation of nuclear and component features within the tissue.

• Nuclear Staining intensity which demonstrates the chromatin detail clearly.


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• Staining colour, intensity and balance may not be consistent across the preparation but

allows clear distinction between nuclear detail and other non-nuclear features.

• The tissue constituents may lack appropriate colour spectrum in some areas.

• Primary and counterstain intensity may be weak or intense but does not obscure detail.


results.

• There may be some uneven staining or background staining but detail is visible in the

majority of the tissue.



• There may be some microtomy or processing irregularities, but the material is adequate to

assess.

• Dehydration, covering and labelling does not impair the visualisation of the tissue and its

components.

• The overall preparation and staining is good and allows full visualisation of the tissue and its

components. Loss of staining or preparation quality is not detrimental to the identification of

tissue constituent details.

Score 3 - Pass Adequate nuclear and tissue constituent staining in a suitable preparation

to allow visualisation of nuclear and component features within the tissue.

• There may be alteration to nuclear and non-nuclear intensity and colour, but most nuclear

detail is visible.

• The tissue constituents do not show full colour spectrum. Nuclear staining may be under or

over stained.

• Primary and counterstain may be weak or intense but detail is still visible. Some background

staining.


results.

• Staining may not be even across the preparation.



• There may be some microtomy or processing irregularities.

• The dehydration, covering and labelling may obscure visualisation of the tissue and its

components but there is sufficient visible to assess.


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Score 2 - Borderline Fail Suboptimal nuclear and or tissue constituent staining which does not

allow full visualisation of nuclear and or component features. The

preparation quality or method does not allow full observation within the

tissue.

• There may be alteration to nuclear and non-nuclear staining intensity and colour, but some

nuclear detail is visible.

• The tissue constituents do not show full colour spectrum. Nuclear and / or tissue component

staining may be under or over stained.

• Primary and counterstain may be intense and / or background staining may obscure detail.

• Demonstration is inappropriate based on the method employed and the expected staining

results.

• Staining may not be even across the preparation.

• The preparation quality may be suboptimal obscuring some tissue detail. There may be

areas of microtomy or processing irregularities.

• The dehydration, covering and labelling may hinder tissue visualisation.

Score 1- Fail The nuclear and or tissue constituent staining does not allow visualisation

of nuclear and / or component features or the preparation quality does

not allow clear observation within the tissue.

• Alteration to nuclear and non-nuclear staining intensity and colour which obscures nuclear

detail.

• Background staining which obscures nuclear and tissue constituent detail.

• Demonstration is inappropriate based on the method employed and the expected staining

results.

• Uneven or patchy staining amounting to loss of nuclear and tissue constituent detail in the

majority of the tissue.

• Preparation is not suitable to allow clear observation of tissue components.

• The preparation quality may be suboptimal obscuring some tissue detail.

• The dehydration, covering and labelling obscures tissue visualisation.

Score 0 - Non submission No slides submitted for assessment or slides returned late without

contacting the Scheme Manager.


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UK NEQAS CPT Stain Repertoire

General Pathology Scheme


Special A Special B

Diastase / PAS Alcian blue / PAS

Elastin / van Gieson Amyloid (method for)

Gram Grocott

Perls’ Prussian blue Haematoxylin / van Gieson

Reticulin ( silver method for) Masson-Fontana

Ziehl-Neelsen Martius-scarlet-blue (MSB)

Copper Associated Protein (method for)

Trichrome (Not HVG)

Neuropathology Scheme


Special A Special B

Diastase / PAS Axonal Swelling (method for)

Elastin / van Gieson Glial fibres (method for)

Gram Myelin (method for)

Perls’ Prussian blue Neurofibrillary tangles

Reticulin ( silver method for) Nissl substance

Ziehl-Neelsen Senile plaques (method for)

For neuropathology, the methods in list B may include Immunocytochemical techniques where that

is the department’s method of choice.

Renal Biopsy Pathology Scheme Muscle Histochemistry Scheme

Haematoxylin and Eosin Haematoxylin and Eosin

Methenamine Silver NADH

PAS/dPAS Gomori

Elastin / van Gieson Cytochrome Oxidase

Non Gynae Diagnostic Cytology Scheme

Staining techniques: Specimen Types:

Papanicolaou Serous Fluid

Romanowsky Head and Neck

Respiratory

Urine

staining criteria handbook - plusnet - special stain...• uk neqas cpt stain repertoire . ......

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