Stains - Acid fast

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Acid fast

Reviewer: Nat Pernick, M.D., PathologyOutlines.com, Inc. (see Reviewers page)
Revised: 20 April 2011, last major update April 2011
Copyright: (c) 2002-2011, PathologyOutlines.com, Inc.

Terminology
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● Acid fast refers to microorganisms whose cell wall has a high lipid content of mycolic acids and long chain fatty acids, which causes them to bind and retain the complex basic dye carbol-fuchsin even after strong decolorization with acid-alcohol (thus “acid-fast”)
● Partially acid fast organisms exhibit both acid fast and non acid fast bacilli and filaments in a single strain

Acid fast organisms
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● Acid fast organisms include Mycobacteria, Cryptosporidium parvum, Isospora and Cyclospora cysts, hooklets of cysticerci
● Partially acid fast organisms include nocardiae, Dietzia (Int J Syst Evol Microbiol 2006;56:1667), Rhodococcus (South Med J 1991;84:1217), Gordonia (Emerg Infect Dis 2000;6:382), Tsukamurella (J Med Case Reports 2008;2:207), rarely Mycobacterium peregrinum (J Clin Microbiol 2005;43:2015)
● Note: nucleic acid based tests can rapidly detect and speciate mycobacteria (Arch Pathol Lab Med 2008;132:1333, Thorax 2008;63:317)

Acid fast methods
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● Ziehl-Neelsen (classic): common method; bacteria stain bright red due to retention of carbol-fuchsin dye; background is methylene blue counterstain; procedure involves heat (#1, #2)
● Ziehl-Neelsen (modified bleach): may be more sensitive than classic stain (Acta Cytol 2008;52:325)
● Kinyoun: common method; uses more concentrated fuchsin dye and lipid solvent, but no heat; bacteria stain bright red against green background (#1, #2)
● Fite: to detect M. leprae (leprosy) and Rhodococcus (Diagn Cytopathol 2001;24:244); combines peanut/vegetable oil with xylene to minimize exposure of bacteria cell wall to organic solvents and protect precarious acid-fastness of organism (#1, #2)
● Ellis and Zabrowarny: protocol excludes phenol; procedure (J Clin Pathol 1993;46:559)
● Auramine-rhodamine: mixture of Auramine O and Rhodamine B dyes, auramine binds to mycolic acid in cell wall; detection requires a fluorescence microscope (mercury vapor lamp or LED), but is the most sensitive stain for mycobacteria (Hum Pathol 1984;15:1085); saves time in searching for microorganisms (Clin Infect Dis 2008;47:203); procedure
● Water filters are recommended to reduce false positives due to nonTB mycobacteria (Appl Environ Microbiol 2007;73:6296)

Micro images - Cryptosporidium
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Oocysts-modified acid-fast stain

Stool specimen (Ziehl-Neelsen)

Oocysts-auramine-rhodamine stain

Micro images - Mycobacterium tuberculosis
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Ziehl-Neelsen stains of site-unknown, cervix and lung (left to right)

Auramine stain of lung

Micro images - Mycobacterium avium complex
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Ziehl-Neelsen stains of site-unknown, breast and colon (left to right)

Ziehl-Neelsen stain of lymph node post bCG vaccination

Micro images - Mycobacterium leprae
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Fite stain of liver

Micro images - Nocardia
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Fite-Faraco Modified acid fast stain of lung

Micro images - Other
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Ziehl-Neelsen with light counterstain shows kayexalate particles at site of aspiration pneumonia

Videos
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● Acid fast procedure

End of Stains > Acid fast

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