Acridine Orange Staining
DESCRIPTION
Acridine orange is a fluorochrome that binds to nucleic acids. Method is used for detecting low numbers of organisms in cerebral
spinal fluid, blood, buffy coat preparations from neonates, and tissue specimens. The stain maybe useful in interpreting purulent
specimens that failed to give clear results with the Gram stain. Other microorganisms are well stained with this method: Mycoplasma,
Pneumocystis, Borrelia, Acanthamoeba, Leishmania, Helicobacter.
REAGENTS
0.2 M sodium acetate buffer (pH 3.75). Add Acridine orange to buffer (0.02 g).


PROCEDURE
1. Dry then fix the smear in methanol for 2 minutes.
2. Stain with Acridine orange for 1 minute.
3. Rinse with distilled water.


RESULTS
Bacteria and yeasts stain bright orange/red, and tissue cells stain black to
yellowishgreen.
REFERENCES
1. Emanuel Goldman and Lorrence H. Green (ed.): Practical handbook of microbiology, 2009, 39-40.
NOTES
Blood cells are not fluorescent; the stain can be useful for screening blood cultures for the growth of microorganisms.
(c) Costin Stoica
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