Bacterial growth studied by flow cytometry

Abstract

The feasibility of flow cytometry for measurements on bacteria has been demonstrated by measurements of DNA-associated fluorescence of Escherichia coli K-12 in various phases of cell growth. Bacteria were stained with a combination of ethidium bromide and mithramycin after fixation in 70% ethanol. Cultures grown to stationary phase accumulated in two peaks representing cells with two and four chromosomes. Qualitatively similar histograms were obtained with cells grown in the presence of chloramphenicol, whereas cells of the temperature sensitive strain E 177 (dnaA) ended up with only one chromosome per cell at the restrictive temperature. The fluorescence intensity of cells with one chromosome was about 10³ times smaller than that of human diploid cells. Instrumental resolution at this level of intensity was CV = 5%, whereas peak widths corresponded to CV = 7–8%. Dyes bound to RNA did not appear to contribute significantly to the fluorescence.