Stains - Enzyme Cytochemistry

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Stains

Enzyme Cytochemistry

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.

General
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● Detects enzymatic activity in cytoplasm
● Advantage over immunocytochemistry is determination of enzyme's intracellular localization and intensity of catalytic activity (for research purposes)
● Flow cytometry and immunocytochemistry are often preferred to determine presence of enzyme molecule (but not catalytic activity or localization)
● Enzyme product unites with coupler, which produces localized color at site of enzyme activity
● Fresh smears are preferred, especially for myeloperoxidase; if not possible, store unstained slides away from light

Methods
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● Simultaneous capture: reagent in incubation medium combines with reaction product - example: diazo method for alkaline phosphatase
● Post-incubation coupling: insoluble reaction product is coupled with a colored or opaque substance - example: Rutenburg and Seligman method for acid phosphatase
● Self-colored substrate reaction: water-soluble dye is made insoluble when enzyme removes a hydrophilic group, leading to colored precipitate at the site of enzyme activity
● Intramolecular rearrangement: produces a colored insoluble precipitate at sites of enzyme activity of an otherwise colorless substrate (University of Iowa)

Uses
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● Used for these enzymes: acid phosphatase, alkaline phosphatase, myeloperoxidase, Sudan black B, nonspecific esterase
● Also: acetyl cholinesterase, adenosine triphosphate, adenylate cyclate, catalase, cytochrome oxidase, 5' nucleotidase, nucleoside diphosphatase, succinic dehydrogenase, thiamine pyrophosphatase

Micro images
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Left: AML-M2, Right: AML-M1 show myeloperoxidase staining (AFIP Third Series, Volume 9)

AML-M0: myeloblasts are negative for myeloperoxidase with positive staining in neutrophil (AFIP Third Series, Volume 9)

AML-M1: numerous myeloblasts have Sudan black B positive granules (AFIP Third Series, Volume 9)

AML-M4: chloroacetate esterase stains neutrophils blue, and nonspecific esterase stains monocytes red-brown (AFIP Third Series, Volume 9)

AML-M4: non-specific esterase

AML-M4: lysozyme

End of Stains > Enzyme Cytochemistry

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