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A laser‐light pulse counting method for automatic and sensitive counting of bacteria stained with acridine orange
Author(s) -
Kroll R.G.,
Pinder A.C.,
Purdy P.W.,
Rodrigues Ubaldina M.
Publication year - 1989
Publication title -
journal of applied bacteriology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 0021-8847
DOI - 10.1111/j.1365-2672.1989.tb02465.x
Subject(s) - acridine orange , fluorescence , optics , photomultiplier , laser , materials science , dye laser , wavelength , photon counting , analytical chemistry (journal) , chemistry , optoelectronics , chromatography , physics , detector , apoptosis , biochemistry
A laser‐light pulse counting method was developed and investigated for its ability automatically to count acridine orange‐stained bacteria and microcolonies of bacteria on the surface of polycarbonate membrane filters. The system consisted of an Argon laser which delivered a 5 μm diameter spot of coherent 488 nm wavelength light which was raster scanned across a 5×2 mm area of the membrane. Fluorescence pulses of orange light ( ca 650 nm) were detected via a dichroic mirror and barrier filter with a photomultiplier tube. For microcolony preparations a good relationship between the number of light pulses and cell density down to below 10 2 /ml was observed, but at lower cell densities counting was not reliable, probably because of fluorescent debris. The method was also able to count single cells, but background auto‐fluorescence and photo‐bleaching produced a high and variable background signal in some samples.