Difference between revisions of "Stains"

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This article deals with '''stains'''. H&E isn't the only stain out there.
[[Image:Corpora amylacea high mag.jpg|thumb|300px|[[Hematoxylin and eosin stain]] of benign [[prostate gland]].]]
This article deals with '''stains'''. The H&E stain is only one of many stains that are used in pathology.


Non-H&E stains are often referred to as '''''special stains'''''.
Non-H&E stains are often referred to as '''''special stains'''''.  


=Where to start...=
=Where to start...=
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==Haematoxylin and eosin stain==
==Haematoxylin and eosin stain==
===General===
*Abbreviated ''H&E''.
*Abbreviated ''H&E''.
*Standard bearer in most pathology departments.
{{Main|Hematoxylin and eosin stain}}
 
===Intepretation===
*Blue (haematoxylin) = nucleus.
*Pink (eosin) = cytoplasm.
 
====Images====
<gallery>
Image:Cecal adenocarcinoma.jpg | Cecal adenocarcinoma. H&E stain. (WC)
Image:Basal cell carcinoma - high mag.jpg | [[Basal cell carcinoma]]. H&E stain. (WC)
</gallery>


==Haematoxylin phyloxin saffron stain==
==Haematoxylin phyloxin saffron stain==
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<gallery>  
<gallery>  
Image:Metanephric_adenoma_high_mag.jpg | Metanephric adenoma - PAS (WC)
Image:Metanephric_adenoma_high_mag.jpg | Metanephric adenoma - PAS (WC)
File:Liver biopsy of glycogen storage disorder PAS positive.jpg | Liver biopsy with glycogen storage disease - PAS (WC)
File:Esophageal candidiasis (2) PAS stain.jpg | Esophageal candidiasis - PAS (WC)
</gallery>
</gallery>


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There are several AFB stains:
There are several AFB stains:
*Ziehl-Neelson stain - used to look for ''[[Mycobacterium tuberculosis]]''.
*Ziehl-Neelson stain - used to look for ''[[Mycobacterium tuberculosis]]''.
*Fite stain - used to look for ''[[Mycobacterium leprae]]''.<ref>URL: [http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/FITES.PDF http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/FITES.PDF]. Accessed on: 19 May 2011.</ref>
*Fite stain - used to look for ''[[Mycobacterium leprae]]''<ref>URL: [http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/FITES.PDF http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/FITES.PDF]. Accessed on: 19 May 2011.</ref> and preferred stain for ''[[Mycobacterium avium complex]]''.<ref name=pmid21327589>{{cite journal |authors=Ilyas S, Youssef D, Chaudhary H, Al-Abbadi MA |title=Myocbacterium-avium intracellulare associated inflammatory pseudotumor of the anterior nasal cavity |journal=Head Neck Pathol |volume=5 |issue=3 |pages=296–301 |date=September 2011 |pmid=21327589 |pmc=3173547 |doi=10.1007/s12105-011-0248-0 |url=}}</ref>
*Auramine-rhodamine stain.
*Auramine-rhodamine stain.


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Note:
Note:
*Thick sections (~10 micrometers) are considered a requirement for the stain to work properly.<ref>URL: [http://www.ihcworld.com/_protocols/special_stains/congo_red_bennhold.htm http://www.ihcworld.com/_protocols/special_stains/congo_red_bennhold.htm]. Accessed on: 26 January 2012.</ref>
*[[Cutting|Thick sections]] (~10 micrometers) are considered a requirement for the stain to work properly.<ref>URL: [http://www.ihcworld.com/_protocols/special_stains/congo_red_bennhold.htm http://www.ihcworld.com/_protocols/special_stains/congo_red_bennhold.htm]. Accessed on: 26 January 2012.</ref>
**If the section is too thin... it doesn't work.
**If the section is too thin... it doesn't work.


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Image:Cerebral_amyloid_angiopathy_-_very_high_mag.jpg | Congo red staining in [[cerebral amyloid angiopathy]]. (WC)
Image:Cerebral_amyloid_angiopathy_-_very_high_mag.jpg | Congo red staining in [[cerebral amyloid angiopathy]]. (WC)
</gallery>
</gallery>
==Thioflavin T stain==
==Thioflavin T stain==
===Use===
===Use===
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<gallery>
<gallery>
Image:Globus_pallidus_and_putamen_-_very_low_mag.jpg | Globus pallidus and putamen - H&E-LFB. (WC)
Image:Globus_pallidus_and_putamen_-_very_low_mag.jpg | Globus pallidus and putamen - H&E-LFB. (WC)
File:LFB_CNS_cortex_supratentorial.jpg | Normal cortex - LFB only. (WC/jensflorian)
File:LFB_CNS_cortex_grey-white_matter_junction.jpg | White-grey matter junction - LFB. (WC/jensflorian)
</gallery>
</gallery>
==Giemsa stain==
==Giemsa stain==
===Use===
===Use===
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===Interpretation===
===Interpretation===
*Tissue is light blue/green.
*Tissue is light blue/green.
*Goblet cells are purple.<ref>URL: [http://www.kennedy.ox.ac.uk/facilities/histology/histology-information http://www.kennedy.ox.ac.uk/facilities/histology/histology-information]. Accessed on: 17 August 2015.</ref>
Image:
*[http://path.upmc.edu/cases/case196/images/figure12.jpg Giemsa - colon (amser.org)].<ref>URL: [http://amser.org/index.php?P=AMSER--ResourceFrame&resourceId=6018 http://amser.org/index.php?P=AMSER--ResourceFrame&resourceId=6018]. Accessed on: 17 August 2015.</ref>


==Reticulin stain==
==Reticulin stain==
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**Sarcomas have reticulin around ''each'' cell.
**Sarcomas have reticulin around ''each'' cell.
**Carcinomas have reticulin around clusters of cells.
**Carcinomas have reticulin around clusters of cells.
*Commonly used in neuropathology.
** In adenoma, reticulin highlights the lost acinar structure of normal pituitary gland.
** Paraganglioma (Zellballen architecture)
** Separating schwannoma (basement membrane around each cell) from meingioma in cerebellopontine angle.
** Separating desmoplastic medulloblastoma from classic/anaplastic forms.


===Interpretation===
===Interpretation===
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Image:Liver_reticulin.jpg | [[Liver]]. Reticulin stain. (WC)
Image:Liver_reticulin.jpg | [[Liver]]. Reticulin stain. (WC)
Image:Hepatic_adenoma_high_mag_reticulin.jpg | [[Hepatic adenoma]]. Reticulin stain. (WC)
Image:Hepatic_adenoma_high_mag_reticulin.jpg | [[Hepatic adenoma]]. Reticulin stain. (WC)
File:Zellballen paraganglioma.jpg | Reticulin stain highlighting the "Zellballen" architecture of paraganglioma. (WC/jensflorian)
File:Desmoplastic medulloblastoma reticulin stain pale island.jpg | Reticulin staina round the "pale islands" of a desmoplastic medulloblastoma. (WC/jensflorian)
</gallery>
</gallery>


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<gallery>
<gallery>
Image:Hemosiderosis_high_mag.jpg | Liver [[hemosiderosis]]. Prussian blue stain. (WC/Nephron)
Image:Hemosiderosis_high_mag.jpg | Liver [[hemosiderosis]]. Prussian blue stain. (WC/Nephron)
File:Siderophage iron stain CSF.jpg | CSF Siderophages in subarachnoid hemorrhage. (WC/jensflorian)
</gallery>
</gallery>


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*Useful for differentiating brown pigments (melanin, lipofuscin, tattoo pigment, hemosiderin).
*Useful for differentiating brown pigments (melanin, lipofuscin, tattoo pigment, hemosiderin).
**Stains lipofuscin.
**Stains lipofuscin.
*Useful to detect demyelinating lesions in the CNS.


Notes:
Notes:
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*Described well by ''vetmed.vt.edu''.<ref>Kluver-Barrera stain. URL:[http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exkluvbarr.htm http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exkluvbarr.htm]. Accessed on: 5 May 2010.</ref>
*Described well by ''vetmed.vt.edu''.<ref>Kluver-Barrera stain. URL:[http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exkluvbarr.htm http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exkluvbarr.htm]. Accessed on: 5 May 2010.</ref>
*DDx of brown pigment: Fontana-Masson (melanin), Prussian blue stain (hemosiderin).
*DDx of brown pigment: Fontana-Masson (melanin), Prussian blue stain (hemosiderin).
<gallery>
File:MS Demyelinisation KB 10x.jpg | Encephalomyelitis disseminata (Klüver-Barrera)
</gallery>


==Oil red O stain==
==Oil red O stain==
===Use===
===Use===
*Stains adipose tissue.
*Stains adipose tissue.
*Corroborate diagnosis of [[lipoid pneumonia]].<ref name=pmid25374742>{{Cite journal  | last1 = Yampara Guarachi | first1 = GI. | last2 = Barbosa Moreira | first2 = V. | last3 = Santos Ferreira | first3 = A. | last4 = Sias | first4 = SM. | last5 = Rodrigues | first5 = CC. | last6 = Teixeira | first6 = GH. | title = Lipoid pneumonia in a gas station attendant. | journal = Case Rep Pulmonol | volume = 2014 | issue =  | pages = 358761 | month =  | year = 2014 | doi = 10.1155/2014/358761 | PMID = 25374742 }}</ref>
*Screen for [[GERD]] - positive staining seen in macrophages from [[BAL]] specimens.<ref name=pmid20466562>{{Cite journal  | last1 = Hopkins | first1 = PM. | last2 = Kermeen | first2 = F. | last3 = Duhig | first3 = E. | last4 = Fletcher | first4 = L. | last5 = Gradwell | first5 = J. | last6 = Whitfield | first6 = L. | last7 = Godinez | first7 = C. | last8 = Musk | first8 = M. | last9 = Chambers | first9 = D. | title = Oil red O stain of alveolar macrophages is an effective screening test for gastroesophageal reflux disease in lung transplant recipients. | journal = J Heart Lung Transplant | volume = 29 | issue = 8 | pages = 859-64 | month = Aug | year = 2010 | doi = 10.1016/j.healun.2010.03.015 | PMID = 20466562 }}</ref>
*Uncommon.
*Uncommon.


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Image:Barrett's_mucosa,_higher_magnification,_Alcian_blue_stain_.jpg | [[Barrett's esophagus|Barrett's type mucosa]]. Alcian blue stain. (WC/AFIP)
Image:Barrett's_mucosa,_higher_magnification,_Alcian_blue_stain_.jpg | [[Barrett's esophagus|Barrett's type mucosa]]. Alcian blue stain. (WC/AFIP)
</gallery>
</gallery>
==Sodium sulphate-alcian blue stain==
:''Sulfated alcian blue'' (abbreviated '''SAB'') redirects here.
===Use===
*Identify [[amyloid]].<ref name=pmid55419>{{Cite journal  | last1 = Pomerance | first1 = A. | last2 = Slavin | first2 = G. | last3 = McWatt | first3 = J. | title = Experience with the sodium sulphate-Alcian Blue stain for amyloid in cardiac pathology. | journal = J Clin Pathol | volume = 29 | issue = 1 | pages = 22-6 | month = Jan | year = 1976 | doi =  | PMID = 55419 }}</ref><ref>URL: [http://www.polyrnd.com/products/reagent-assembly-kits/conventional/amyloid-stain---sulfated-alcian-blue-(sab).aspx http://www.polyrnd.com/products/reagent-assembly-kits/conventional/amyloid-stain---sulfated-alcian-blue-(sab).aspx]. Accessed on: October 15, 2014.</ref>
*Less [[specificity|specific]] than [[congo red stain|congo red]] but equally [[sensitivity|sensitive]].
*Does not require polarized light.
===Interpretation===
*Green = amyloid.<ref name=pmid55419/>
**Other things that are green: [[mast cell]]s, mucoid degeneration, basophilic myofibre degeneration, califications.
*Yellow = background.
Image:
*[http://www.ncbi.nlm.nih.gov/pmc/articles/PMC475937/figure/F1/ SAB (nih.gov)].


==Movat's stain==
==Movat's stain==
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*Red = muscle (smooth muscle actin).
*Red = muscle (smooth muscle actin).
*Baby blue = collagen.
*Baby blue = collagen.
*Blue gray = [[amyloid]].<ref name=pmid33002919>{{cite journal |authors=Kunnath-Velayudhan S, Larsen BT, Coley SM, De Michele S, Santoriello D, Colby TV, Bhagat G, Saqi A |title=Masson Trichrome and Sulfated Alcian Blue Stains Distinguish Light Chain Deposition Disease From Amyloidosis in the Lung |journal=Am J Surg Pathol |volume=45 |issue=3 |pages=405–413 |date=March 2021 |pmid=33002919 |doi=10.1097/PAS.0000000000001593 |url=}}</ref>


Notes: <Ref>ULR: [http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/MASSONS.PDF http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/MASSONS.PDF]. Accessed on: 2 November 2011.</ref>
Notes: <Ref>ULR: [http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/MASSONS.PDF http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/MASSONS.PDF]. Accessed on: 2 November 2011.</ref>
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==Hale's colloidal iron stain==
==Hale's colloidal iron stain==
===Use===
*[[Chromophobe renal cell carcinoma]] vs. [[renal oncocytoma]] - chromophobe [[renal cell carcinoma]] +ve.<ref name=pmid9537468>{{cite journal |author=Tickoo SK, Amin MB, Zarbo RJ |title=Colloidal iron staining in renal epithelial neoplasms, including chromophobe renal cell carcinoma: emphasis on technique and patterns of staining |journal=Am. J. Surg. Pathol. |volume=22 |issue=4 |pages=419–24 |year=1998 |month=April |pmid=9537468 |doi= |url=http://meta.wkhealth.com/pt/pt-core/template-journal/lwwgateway/media/landingpage.htm?issn=0147-5185&volume=22&issue=4&spage=419}}</ref>


Notes:
{{Main|Hale's colloidal iron stain}}


===Interpretation===
==von Kossa stain==
*Blue (granular cytoplasmic) staining is positive.<ref name=Ref_WMSP682>{{Ref WMSP|682}}</ref>
===General===
*Type of silver stain.<ref name=pmid8360080>{{Cite journal  | last1 = Rungby | first1 = J. | last2 = Kassem | first2 = M. | last3 = Eriksen | first3 = EF. | last4 = Danscher | first4 = G. | title = The von Kossa reaction for calcium deposits: silver lactate staining increases sensitivity and reduces background. | journal = Histochem J | volume = 25 | issue = 6 | pages = 446-51 | month = Jun | year = 1993 | doi =  | PMID = 8360080 }}</ref>


Images:
*[http://www.ultrapath.org/oldsite/cases99/sep99/images/figure-3.jpg ChRCC Hale's colloidal iron (ultrapath.org)].<ref>URL: [http://www.ultrapath.org/oldsite/cases99/sep99/cotm9-2.html http://www.ultrapath.org/oldsite/cases99/sep99/cotm9-2.html]. Accessed on: 9 October 2011.</ref>
*[http://www.diagnosticpathology.org/content/4/1/21/figure/f1?highres=n ChRCC Hale's colloidal iron (diagnosticpathology.org)].
Notes:
*Often described as a "fastidious" (difficult/demanding) stain.<ref>URL: [http://www.merriam-webster.com/dictionary/fastidious?show=0&t=1319550566 http://www.merriam-webster.com/dictionary/fastidious?show=0&t=1319550566]. Accessed on: 25 October 2011.</ref>
**A few staff think this is a totally useless stain.<ref>ALS. On several occasions in 2009.</ref>
==von Kossa stain==
===Use===
===Use===
*Look for calcium.
*Look for calcium.
*Actually stains phosphates and carbonates as a surrogate for calcium.


===Interpretation===
===Interpretation===
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*[[Cytopathology]].
*[[Cytopathology]].
*[[Helicobacter gastritis]] - organisms are dark blue against a light blue background.<ref>URL: [http://www.ihcworld.com/_protocols/special_stains/diff_quick_ellis.htm http://www.ihcworld.com/_protocols/special_stains/diff_quick_ellis.htm]. Accessed on: 30 August 2012.</ref>
*[[Helicobacter gastritis]] - organisms are dark blue against a light blue background.<ref>URL: [http://www.ihcworld.com/_protocols/special_stains/diff_quick_ellis.htm http://www.ihcworld.com/_protocols/special_stains/diff_quick_ellis.htm]. Accessed on: 30 August 2012.</ref>
====Images====
<gallery>
Image: Lung adenocarcinoma - Diff-Quik -- high mag.jpg | [[Pulmonary_cytopathology#Adenocarcinoma|Lung adenocarcinoma]] - DQ - high mag. (WC)
Image: Lung adenocarcinoma - Diff-Quik -- very high mag.jpg | Lung adenocarcinoma - DQ - very high mag. (WC)
Image: Lung small cell carcinoma - Diff-Quik -- very high mag.jpg | Lung SmCC - DQ - very high mag. (WC)
Image: Lung small cell carcinoma - Diff-Quik -- extremely high mag.jpg | Lung SmCC - DQ - extremely high mag. (WC)
</gallery>


===Wright stain===
===Wright stain===
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==Papanicolaou stain==
==Papanicolaou stain==
*Abbreviated ''Pap stain''.
*Abbreviated ''Pap stain''.
*Can be thought of as the [[H&E stain|H&E]] of [[cytopathology]].
{{Main|Papanicolaou stain}}
**It is a modified [[H&E stain]].
*Specimens are fixed in ethanol.
*Good for seeing nuclear detail.
*Out-of-focus cytoplasm is translucent; allows one to focus overlapped cells in different planes.
 
===Use===
*Cytopathology.
 
===Interpretation===
*Blue/purple = nucleus.
*Green/pink = cytoplasm.
*Orange = keratin.
 
====Image====
<gallery>
Image:Urine_citology_urothelial_carcinoma_2.jpg | Pap stain - [[urine cytology]] (WC)
</gallery>


==Fontana-Masson stain==  
==Fontana-Masson stain==  
*[[AKA]] ''Masson-Fontana stain'',<ref name=pmid16081962>{{Cite journal  | last1 = Gaitanis | first1 = G. | last2 = Chasapi | first2 = V. | last3 = Velegraki | first3 = A. | title = Novel application of the masson-fontana stain for demonstrating Malassezia species melanin-like pigment production in vitro and in clinical specimens. | journal = J Clin Microbiol | volume = 43 | issue = 8 | pages = 4147-51 | month = Aug | year = 2005 | doi = 10.1128/JCM.43.8.4147-4151.2005 | PMID = 16081962 }}</ref> ''Fontana-Masson stain for melanin'', ''melanin stain''.
*[[AKA]] ''Masson-Fontana stain'',<ref name=pmid16081962>{{Cite journal  | last1 = Gaitanis | first1 = G. | last2 = Chasapi | first2 = V. | last3 = Velegraki | first3 = A. | title = Novel application of the masson-fontana stain for demonstrating Malassezia species melanin-like pigment production in vitro and in clinical specimens. | journal = J Clin Microbiol | volume = 43 | issue = 8 | pages = 4147-51 | month = Aug | year = 2005 | doi = 10.1128/JCM.43.8.4147-4151.2005 | PMID = 16081962 }}</ref> ''Fontana-Masson stain for melanin'', ''melanin stain''.
*A type of silver stain.
{{Main|Fontana-Masson stain}}
 
Stains:
#Melanin.
#"Argentaffin granules" of the digestive tract.
#Pigment deposition due to minocycline treatment.<ref name=pmid15605966>{{Cite journal  | last1 = Patterson | first1 = JW. | last2 = Wilson | first2 = B. | last3 = Wick | first3 = MR. | last4 = Heath | first4 = C. | title = Hyperpigmented scar due to minocycline therapy. | journal = Cutis | volume = 74 | issue = 5 | pages = 293-8 | month = Nov | year = 2004 | doi =  | PMID = 15605966 }}</ref>
 
===Use===
*Stain for melanin.
**Used to differentiate brown pigments (lipofuscin, hemosiderin, melanin).<ref>URL: [http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exfontana.htm http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exfontana.htm]. Accessed on: 5 May 2010.</ref>
***Many pathologists prefer IHC, i.e. ''Melan A'' over this stain.
 
Image:
*[http://education.vetmed.vt.edu/curriculum/VM8054/labs/Lab2/Examples/exfontana.htm Fontana-Masson stain (vetmed.vt.edu)].


==Schmorl's stain==
==Schmorl's stain==
Line 711: Line 711:


Image:
Image:
*[http://www.bris.ac.uk/vetpath/cpl/msb.jpg MSB (bris.ac.uk)].
*[https://www.ole.bris.ac.uk/bbcswebdav/institution/Faculty%20of%20Health%20Sciences/Veterinary%20Science/eLearning%20resources/Pathology%20Laboratory%20Protocols/hst/msb.html MSB (bris.ac.uk)].


Ref.:<ref>URL: [http://www.bris.ac.uk/vetpath/cpl/msb.html http://www.bris.ac.uk/vetpath/cpl/msb.html]. Accessed on: 26 November 2010.</ref>
Ref.:<ref>URL: [http://www.bris.ac.uk/vetpath/cpl/msb.html http://www.bris.ac.uk/vetpath/cpl/msb.html]. Accessed on: 26 November 2010.</ref>
Line 740: Line 740:
*Collagen = bright red.
*Collagen = bright red.
*Muscle = dull red.
*Muscle = dull red.
<gallery>
File:Cerebral aneurysm EVG stain.jpg |EVG stain of a cerebral aneurysm. (WC/jensflorian)
</gallery>


==Copper stain==
==Copper stain==
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===General===
===General===
*Used in [[medical liver disease|medical liver]] biopsies - used to find copper.  
*Used in [[medical liver disease|medical liver]] biopsies - stains sulfhydrl groups and identifies:
**Copper-associated protein.
**Elastin.
**[[Hepatitis B]] surface antigen.<ref name=pmid7822848>{{Cite journal  | last1 = Ghosh | first1 = AK. | last2 = Dasgupta | first2 = A. | last3 = Raha | first3 = K. | last4 = Jana | first4 = A. | last5 = Majumdar | first5 = DN. | title = Hepatic histology in chronic liver disease in hepatitis B surface antigen positive cases. | journal = J Indian Med Assoc | volume = 92 | issue = 10 | pages = 333-5 | month = Oct | year = 1994 | doi =  | PMID = 7822848 }}</ref>


===Interpretation===
===Interpretation===
Features:<ref>URL: [http://www.nottingham.ac.uk/pathology/protocols/shikata.html http://www.nottingham.ac.uk/pathology/protocols/shikata.html]. Accessed on: 24 January 2011.</ref>
Features:<ref>URL: [http://www.nottingham.ac.uk/pathology/protocols/shikata.html http://www.nottingham.ac.uk/pathology/protocols/shikata.html]. Accessed on: 24 January 2011.</ref>
*Purple/brown = elastin fibres.
*Dark purple/brown = elastin fibres, HBsAg and copper-associated protein
*Light purple = background
*Red = nuclei (only if counter-stain used)
 
==Gömöri Trichrome stain==
*Named after George Gömöri<ref>GOMORI, G. - A rapid one-step trichrome stain. Am. J. Clin. Path. 20: 661-664, 1950.</ref>
 
===General===
*Used in [[muscle biopsies]] - used to find abnormal mitochondrial deposits.
 
===Interpretation===
*Dark green = muscle fibers.
*Red = nuclei.
*Red = nuclei.
*Light purple = background
*Bright red = mitochondria, red blood cells.
*??? = Copper associated protein.
 
Images:
<gallery>
File:Ragged red fibers in MELAS.jpg | Ragged red fibers in MELAS, a mitochondrial disease. (WC)
File:Dilated peri-tubular capillaries filled with sickled RBCs, original Gomori's trichrome stain.jpg | Sickle cell nephropathy. (WC)
</gallery>
 
==Miller stain==
===General===
*Stains elastin.
*Used to identify blood vessels and [[pleural invasion]] in [[lung cancer]].
 
===Interpretation===
Staining:<ref>URL: [https://www.ihcworld.com/_protocols/special_stains/miller's_elastic_ellis.htm https://www.ihcworld.com/_protocols/special_stains/miller's_elastic_ellis.htm]. Accessed on: 28 August 2015.</ref>
*Black = elastin fibres, granules in mast cells.
*Red = collagen.
*Yellow = muscle, fibrin, [[erythrocytes]].
*Green/brown = nuclei.
 
====Images====
<gallery>
Image: Normal visceral pleura of lung - Miller -- high mag.jpg | Miller stain showing lung tissue. (WC)
</gallery>


=See also=
=See also=

Latest revision as of 16:05, 1 May 2024

This article deals with stains. The H&E stain is only one of many stains that are used in pathology.

Non-H&E stains are often referred to as special stains.

Where to start...

Principles

When considering additional (i.e. special) stains one should (in order) do the following:[1]

  1. Make sure one has exhausted the clinical history; history is considered the best special stain.
  2. Special stains (below).
  3. Immunohistochemistry (dealt with in a separate article).
  4. Molecular testing, electron microscopy.

Common stains

  1. H&E stain.
  2. PAS stain.
  3. PAS-D stain.
  4. AFB stains, e.g. Ziehl-Neelsen stain.
  5. Congo red.
  6. GMS stain.
  7. Gram stain.

Immunohistochemistry

General

  • Abbreviated IHC.

Interpretation

Simple version:

  • Positive is (usually): brown.
  • Negative tissue is: light blue.

Important notes:

  • One has to know where the target (of the antibody) is supposed to be, i.e. cytoplasm vs. cell membrane.
  • The edge of the tissue may have light staining - edge effect.
  • If everything is brown... suspect that it didn't work.
  • In some situations you're blessed with an internal control, e.g. in renal tumours CD10 will stain RCC and the proximal tubule, in GISTs - CD117 the mast cells are positive.

Work-up of infection

It often not possible to be definitive by staining.[2]

Basic panel:

  • Gram stain - for bacteria.
  • GMS stain - fungal stain.
  • PAS (or PAS-D) - fungal stain.

Fungi

Fungi are a type of microorganisms. They are seen by pathologist every once in a while.

Specific stains

What follows is a big list... of stains.

Haematoxylin and eosin stain

  • Abbreviated H&E.

Haematoxylin phyloxin saffron stain

General

  • Abbreviated HPS.
  • An alternative to the H&E stain - some pathol. departments use this as their standard.

Interpretation

  • Haematoxylin = blue -- stains nucleus.
  • Phyloxin = pink -- stains muscle and cytoplasm.
  • Saffron = yellow -- stains collagen.
  • An alternative to H&E stain.
    • Fibrosis is easier to see on HPS than H&E... as one can see the collagen.

Images

Periodic acid Schiff stain

  • Abbreviated PAS.

Primary application

  • Kidney biopsies, medical.
  • Liver biopsies, medical.
    • Positive in alpha-1 antitrypsin deficiency.

Utility

  • Stains - lipofuscin,[3] basement membranes, fungi, glycogen, (neutral) mucin.

Interpretation

  • Magenta = glycogen, mucin, fungi.
  • Blue = nuclei.

Ref.:[4]

Image

Periodic acid Schiff fungal stain

  • Abbreviated PASF.

Primary application

Interpretation

  • Light purple = fungi.
  • Light green = background.
  • Washed-out light purple = Gram positive bacilli.

Note:

  • This is much improved over the PAS in the context of skin, as the background is similar to the fungal organisms.

Periodic acid Schiff with diastase

  • Abbreviated: PAS-D and PASD.

General

  • Diastase digests the glycogen.
  • "PAS diastase resistant"[5] implies PASD +ve and PAS +ve.

Use

  • Stains mucin.
  • Used to identify glycogen (together with PAS stain).
    • Glycogen = clear (digested) on PAS-D.
    • Glycogen = magenta on PAS.

Notes: [6]

Interpretation

  • Light purple = fungi.
  • Light blue/pink = background. ???

Gomori methenamine-silver stain

  • Abbreviated GMS.

Note:

  • GMS is "Grocott's methenamine Silver" according to WMSP.[7]

Use

Image

Acid-fast bacilli stains

  • Abbreviated: AFB.

There are several AFB stains:

Ziehl-Neelsen stain

  • Most popular acid-fast bacilli stain.
  • Stains other mycobacteria -- not specific for tuberculosis.
    • Stains Nocardia.[10]

Image

Fite stain

Interpretation:

  • Red = AFB.
  • Blue = background.

Auramine-rhodamine stain

  • Fluorescent stain.

Image

Kinyoun stain

Congo red stain

Use

  • Used to look for amyloid.
    • Mnemonic: CRAP = congo red amyloid protein.
  • An alternate stain for amyloid is Thioflavin T.

Note:

  • Thick sections (~10 micrometers) are considered a requirement for the stain to work properly.[13]
    • If the section is too thin... it doesn't work.

Interpretation

  • Amyloid = pink/red.
  • Nuclei = blue.

Ref.:[14]

Image

Thioflavin T stain

Use

Interpretation

  • Amyloid = green.

Image: Amyloid (inano.au.dk).

Gram stain

Use

  • "It is useless for finding bacteria."[15]
    • If they are to be seen... they'll be visible on H&E.

Note:

  • Microbiology is better at finding organisms than pathology.
    • They have one significant advantage -- if a small amount of bugs are present... they grows into a large (obviously visible) colony.

DDx for common patterns

A short list of bacteria and their characteristics:[16]

Shape\Gram stain Positive Negative Variable or negative
Bacilli Clostridium difficile, Bacillus anthracis, Nocardia spp. Escherichia coli, Helicobacter pylori, Yersinia pestis, Hemophilus influenzae Mycobacterium tuberulosis, Legionella pneumophila[17]
Cocci Streptococcus pneumoniae, Staphylococcus aureus Neisseria meningitidis, Moraxella catarrhalis

Interpretation

  • Purple (or blue) = Gram positive organisms.
  • Red = Gram negative organisms, nuclei.[18]
  • Yellow = background.

Notes:

  • Many of the bacteria are quite small relative to lymphocytes; Escherichia coli is 1-2 micrometers long x 0.25 micrometers in diameter.[19]
  • Epithelial cell nuclei & stromal cell nuclei may stain red.
  • Memory device: purple = positive.

Images

Luxol fast blue stain

  • Abbreviated LFB.

Use

  • Neuropathology, myelin stain.

Intepretation

  • Blue = myelinated fibers (contain lipoproteins), lipofuscin.[20]
    • Lack of blue (where it ought to be) = demyelination.
  • Purple = nerve cell (e.g. neuron).
  • Neutrophils = pink.

Ref.:[21]

Image

Giemsa stain

Use

Interpretation

  • Tissue is light blue/green.
  • Goblet cells are purple.[23]

Image:

Reticulin stain

Use

  • Liver biopsy, medical.
    • Demonstrates the reticular fibers (in cirrhosis the fibers are disrupted).
  • Before IHC, reticulin was used to differentiate sarcomas from carcinomas:[25]
    • Sarcomas have reticulin around each cell.
    • Carcinomas have reticulin around clusters of cells.
  • Commonly used in neuropathology.
    • In adenoma, reticulin highlights the lost acinar structure of normal pituitary gland.
    • Paraganglioma (Zellballen architecture)
    • Separating schwannoma (basement membrane around each cell) from meingioma in cerebellopontine angle.
    • Separating desmoplastic medulloblastoma from classic/anaplastic forms.


Interpretation

  • Black = reticular fibers.
  • Red = nuclei.

Notes:[26]

Images

Cresyl violet stain

Use

  • Used at some places (e.g. SMH) to look for Helicobacter organisms.

Interpretation

  • Everything is shades of blue.
    • Helicobacter stains blue.

Prussian blue stain

  • AKA Perl's iron stain.

Use

  • Useful for iron and hemosiderin; useful for differentiating brown pigments (melanin, lipofuscin, tattoo pigment, hemosiderin).

Interpretation

  • Blue = iron.

Image:

Notes:

  • Described well by vetmed.vt.edu.[27]
  • DDx of brown pigment: Fontana-Masson (melanin), Kluver-Barrera stain (lipofuscin).

Images

Kluver-Barrera stain

Combination of:

  • Luxol Fast Blue,
  • Cresyl Violet,
  • Special component for lipofuscin.

Use

  • Useful for differentiating brown pigments (melanin, lipofuscin, tattoo pigment, hemosiderin).
    • Stains lipofuscin.
  • Useful to detect demyelinating lesions in the CNS.

Notes:

  • PAS also stains lipofuscin and is more commonly available.

Interpretation

  • Blue pigmented granules = lipofuscin.

Notes:

  • Described well by vetmed.vt.edu.[28]
  • DDx of brown pigment: Fontana-Masson (melanin), Prussian blue stain (hemosiderin).

Oil red O stain

Use

  • Stains adipose tissue.
  • Corroborate diagnosis of lipoid pneumonia.[29]
  • Screen for GERD - positive staining seen in macrophages from BAL specimens.[30]
  • Uncommon.

Notes:

  • Must be done on fresh tissue, i.e. it cannot be fixed in formalin.

Interpretation

  • Red = fat.

Images

Warthin-Starry stain

Background:

  • Developed by a bunch of pathologists in Michigan to look for spirochetes.[31]

Use

  • Find spirochetes, e.g. syphilis (Treponema pallidum),[32] cat-scratch disease (Bartonella henselae).
  • Find Helicobacter spp., e.g. Helicobacter pylori -- Mount Sinai Hospital.[33]

Interpretation:[34]

  • Spirochetes - black.
  • Background - yellow.

Image

Notes:

  • Considered a "dirty" stain - picks-up junk in the background.[35]

Dieterle stain

Considered a variant of the Steiner stain.[36]

Use

  • Find spirochetes, e.g. syphilis (Treponema pallidum),[37] donovan bodies (leishmaniasis),[38] Helicobacter pylori and Bartonella henselae (Cat-scratch disease).[39]

Interpretation

  • Spirochetes - black.
  • Background - yellow.

Images

www:

Bielschowsky stain

Abbreviated: Biel stain.

Use

  • Stains glial tissue, i.e. brain.
    • Demonstrates neurofibrillary tangles, senile plaques (as in Alzheimer's disease).

Interpretation

  • Black = axons, tangles, plaques.
  • Brown/dark brown = plaque, vascular amyloid.
  • Yellow/brown = other.

Ref.: [40]

Image

Mucicarmine stain

  • Stains some mucins... uses the dye carmine.

Use

  • Identify mucin.
  • Malignant cells that produce mucin... carcinomas.[41]

Interpretation

  • Carmine with metanil yellow and Weigert's Hematoxylin:[42]
    • Blue/black = nucleus.
    • Yellow = background.
    • Red = mucin.[43]

Images

www:

Alcian blue stain

General

  • Stains acidic mucin (pH=2.5); Alcian blue = Acidic.
    • A variant uses pH=1.0.[7]

Note:

  • Alcian blue (not otherwise specified) usu. refers to the pH=2.5.[44]

Use

Note:

  • Esophageal submucosal glands - alcian blue positive.

Interpretation

  • Blue = acidic mucins.[45]

Notes:

Image

Sodium sulphate-alcian blue stain

Sulfated alcian blue (abbreviated 'SAB) redirects here.

Use

Interpretation

  • Green = amyloid.[46]
    • Other things that are green: mast cells, mucoid degeneration, basophilic myofibre degeneration, califications.
  • Yellow = background.

Image:

Movat's stain

Use

Components

  • Acid fuchsin, alcian blue, crocein scarlet, elastic hematoxylin, and saffron.[48][49]

Interpretation of Movat stain

  • Black = nuclei and elastic fibers.
  • Yellow = collagen and reticular fibers.
  • Blue = mucin, ground substance.
  • Red (intense) = fibrin.
  • Red = muscle.

Reference: [49]

How to remember? A.: Primary colours (red, blue, yellow) + black.

Images

Masson's trichrome stain

General

  • Collagen vs. muscle.

Interpretation

  • Black = nuclei.
  • Red = muscle (smooth muscle actin).
  • Baby blue = collagen.
  • Blue gray = amyloid.[50]

Notes: [51]

Elastic trichrome stain

General:

  • "Elastic trichrome" is one important variant of Masson's trichrome.

Interpretation - as above in Masson's trichrome - plus:

  • Black = nuclei and elastin.

Mallory trichome stain

General

  • Collagen vs. muscle.
  • May be done with elastin.

Site

  • Kidney Bx (to assess for fibrosis).
  • Liver Bx (to assess for cirrhosis).
  • Cardiovascular/lung (to see differentiate the layers of the arteries, and arteries from veins).

Interpretation

  • Black = nuclei.
  • Red = muscle (smooth muscle actin).
  • Green = collagen.

Image

Haematoxylin orcein phyloxin saffron stain

  • Abbreviated HOPS.[52]
  • It should not be confused with the HPS stain.

Interpretation

  • Blue (haematoxylin) = nuclei.
  • Black (orcein) = elastin.
  • Red (phyloxin) = muscle.
  • Yellow (saffron) = collagen.

Jones stain

  • AKA PAS methenamine technique.[53]
  • AKA Methenamine PAS, abbreviated MPAS.

Use

  • Visualize basement membrane in kidney biopsies.

Interpretation

  • Black = basement membrane.
  • Blue = nuclei.
  • Pink = other structures/background.

Notes:[54]

Images

Hale's colloidal iron stain

von Kossa stain

General

  • Type of silver stain.[55]

Use

  • Look for calcium.
  • Actually stains phosphates and carbonates as a surrogate for calcium.

Interpretation

  • Black = calcium.[7]

Toluidine blue stain

Use

Interpretation

  • Dark blue - nuclei, mast cell granules (darker than nuclei).
  • Light blue - cytoplasm.
  • Red/magneta - cartilage. (???)

Refs: looks a bit sketchy[58], [59]

Image

www:

Romanowsky stain

  • Occasionally spelled Romanowski.
  • Many variants of this stain exist.
  • Specimens are air-dried.

Interpretation:[60]

  • Red - RBCs, eosinophil granules.
  • Blue (basophilic) - lymphocyte cytoplasm.
  • Purple - nuclear chromatin, neutrophil granules, platelets.

Field stain

  • Variant of the Romanowsky stain for rapid processing.
  • Tends to "blow-up" cell, i.e. cells are larger vis-a-vis Pap stain.

Diff-Quik

Pronounced Diff-Quick.
  • Proprietary variant of Romanowsky stain.[61]

Uses:

Images

Wright stain

  • A variant of the Romanowsky stain; popular in North American.

Use:

  • Blood films.

May-Grünwald-Giemsa stain

  • A variant of the Romanowsky stain; popular in Europe.
  • Abbreviated MGG.

Use:

  • Blood films.
  • Cytopathology.

Papanicolaou stain

  • Abbreviated Pap stain.

Fontana-Masson stain

  • AKA Masson-Fontana stain,[63] Fontana-Masson stain for melanin, melanin stain.

Schmorl's stain

  • Stains melanin.
    • Similar to Fontana-Masson stain.

Notes:[64]

Martius scarlet blue stain

General

  • Stains connective tissue and fibrin.[65]
  • Abbreviated MSB.

Use:

Interpretation

  • Muscle and fibrin - red.
  • Nuclei = brown/black.
  • Collagen - blue.
  • Red blood cells - yellow.

Image:

Ref.:[66]

Picro-Mallory stain

General

  • Find fibrin.

Interpretation[67]

  • Fibrin = red.
  • Erythrocytes = yellow.
  • Connective tissue = blue.

Image:

Verhoeff-van Gieson stain

Verhoeff stain redirects here.
  • AKA Elastic van Gieson stain, abbreviated EVG.

General

  • Similar to Masson Trichrome & Verhoeff stain.[68]

Use:

  • Examine large blood vessels.[69]

Interpretation

  • Elastin = black.
  • Collagen = bright red.
  • Muscle = dull red.

Copper stain

General

Note:

  • Copper staining is a non-specific finding seen in many liver diseases; it is associated with impaired bile secretion.[70]

Interpretation

  • Copper = red granules.

Images:

Shikata stain

  • AKA Orcein stain for copper-protein.
  • AKA Shikata-Cu,[72]
  • AKA Shikata's orcein staining.[73]

General

  • Used in medical liver biopsies - stains sulfhydrl groups and identifies:

Interpretation

Features:[75]

  • Dark purple/brown = elastin fibres, HBsAg and copper-associated protein
  • Light purple = background
  • Red = nuclei (only if counter-stain used)

Gömöri Trichrome stain

  • Named after George Gömöri[76]

General

Interpretation

  • Dark green = muscle fibers.
  • Red = nuclei.
  • Bright red = mitochondria, red blood cells.

Images:

Miller stain

General

Interpretation

Staining:[77]

  • Black = elastin fibres, granules in mast cells.
  • Red = collagen.
  • Yellow = muscle, fibrin, erythrocytes.
  • Green/brown = nuclei.

Images

See also

References

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  2. Woods GL, Walker DH (July 1996). "Detection of infection or infectious agents by use of cytologic and histologic stains". Clin. Microbiol. Rev. 9 (3): 382-404. PMC 172900. PMID 8809467. http://cmr.asm.org/cgi/pmidlookup?view=long&pmid=8809467.
  3. Kovi J, Leifer C (July 1970). "Lipofuscin pigment accumulation in spontaneous mammary carcinoma of A/Jax mouse". J Natl Med Assoc 62 (4): 287–90. PMC 2611776. PMID 5463681. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2611776/pdf/jnma00512-0077.pdf.
  4. http://library.med.utah.edu/WebPath/HISTHTML/MANUALS/PAS.PDF
  5. Qizilbash, A.; Young-Pong, O. (Jun 1983). "Alpha 1 antitrypsin liver disease differential diagnosis of PAS-positive, diastase-resistant globules in liver cells.". Am J Clin Pathol 79 (6): 697-702. PMID 6189389.
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