DESCRIPTION
Method for staining flagella.
REAGENTS
Solution 1 (mordant): mix 25 ml of a saturated aqueous solution of aluminum potassium sulfate, 50 ml of 10% tannic acid solution,
and 5 ml of a 5% ferric chloride solution. The resulting mixture turns black.
Solution 2 (stain): slowly add concentrated ammonium hydroxide (2-4 ml) to 90 ml
of a 5% silver nitrate solution until the brown precipitate formed just redissolves. Then add more 5% silver nitrate drop by drop to the
solution until a faint cloudiness persists; this takes from 2 to 20 ml.
Both staining solutions are stable for at least 6 months if kept in the dark at 5°C.
PROCEDURE
Put on slide 4 inoculating loopfuls of sterile distilled water then pick up an isolated colony from agar plate and homogenize. Dry the
slide.
Cover the smear with solution 1 (mordant) for 4 min. Then rinse gently with distilled water and cover the slide with solution 2 (stain).
Apply heat until steam is formed, and wait for 4 min. Rinse the slide gently with distilled water while held horizontally to float off any
loose precipitate and then air dry it.
RESULTS
Cells and flagella are black-colored. The flagella should be clearly seen to originate from the rounded polar regions or from the lateral
side of the rods.
NOTES
Better results are obtained from liquid and semi-solid media than from agar plates.
Poor staining results are obtained from mucoid strains.
REFERENCES
Method for staining flagella.
REAGENTS
Solution 1 (mordant): mix 25 ml of a saturated aqueous solution of aluminum potassium sulfate, 50 ml of 10% tannic acid solution,
and 5 ml of a 5% ferric chloride solution. The resulting mixture turns black.
Solution 2 (stain): slowly add concentrated ammonium hydroxide (2-4 ml) to 90 ml
of a 5% silver nitrate solution until the brown precipitate formed just redissolves. Then add more 5% silver nitrate drop by drop to the
solution until a faint cloudiness persists; this takes from 2 to 20 ml.
Both staining solutions are stable for at least 6 months if kept in the dark at 5°C.
PROCEDURE
Put on slide 4 inoculating loopfuls of sterile distilled water then pick up an isolated colony from agar plate and homogenize. Dry the
slide.
Cover the smear with solution 1 (mordant) for 4 min. Then rinse gently with distilled water and cover the slide with solution 2 (stain).
Apply heat until steam is formed, and wait for 4 min. Rinse the slide gently with distilled water while held horizontally to float off any
loose precipitate and then air dry it.
RESULTS
Cells and flagella are black-colored. The flagella should be clearly seen to originate from the rounded polar regions or from the lateral
side of the rods.
NOTES
Better results are obtained from liquid and semi-solid media than from agar plates.
Poor staining results are obtained from mucoid strains.
REFERENCES
- Rhodes, M. E. 1958. The cytology of Pseudomonas spp. as revealed by a silver-plating staining method. J. Gen. Microbiol. 18:
639-648. - West M, Burdash NM, Freimuth F. 1977. Simplified silver-plating stain for flagella. J Clin Microbiol 6:.https://doi.org/10.1128/jcm.
6.4.414-419.1977
| Modified Rhodes' silver-plating technique |
(c) Costin Stoica
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