Open AccessAn investigation of staining methods to determine total cell numbers and the number of respiring micro‐organisms in samples obtained from the field and the laboratory
Author(s) -
Swannell Richard P.J.,
Williamson Francis A.
Publication year - 1988
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1988.tb02698.x
Subject(s) - formazan , ethidium bromide , malachite green , staining , glutaraldehyde , dapi , dichlorofluorescein , cell counting , bromide , tetrazolium chloride , fluorescence , chromatography , differential staining , chemistry , colorimetry , biology , microbiology and biotechnology , cell , biochemistry , organic chemistry , medicine , dna , physics , ischemia , adsorption , quantum mechanics , cell cycle , cardiology , intracellular , genetics
Dyes were evaluated in combination with 2‐(4‐iodophenyl)‐3‐(4‐nitrophenyl)‐5‐phenyl tetrazolium chloride (INT) to enable total cell numbers and the numbers of respiring cells to be determined on the same preparation. Malachite green and 4′,6‐diamidino‐2‐phenylindole (DAPI) were unsuitable counter‐stains. Cells which contained INT formazan crystals could be stained with ethidium bromide or auramine. At high concentrations of INT formazan, auramine fluorescence was reduced, although this effect was partially rectified by prior fixation with glutaraldehyde. Staining with ethidium bromide produced a strong fluorescence in cells containing crystals of INT formazan. This observation was developed into a procedure which allowed total cells to be determined and provided a useful estimate of the number of respiring cells in samples obtained from the laboratory and the field.