Physical characterization and quantification of bacteria by sedimentation field-flow fractionation

Abstract

Studies in microbial ecology require accurate measures of cell number and biomass. Although epifluorescence microscopy is an accepted and dependable method for determining cell numbers, current methods of converting biovolume to biomass are error prone, tedious, and labor-intensive. This paper describes a technique with sedimentation field-flow fractionation to enumerate bacteria and determine their density, size, and mass. Using cultured cells of different shapes and sizes, we determined optimum values for separation run parameters and sample-handling procedures. The technique described can separate and detect 4',6-diamidino-2-phenylindole-stained cells and generate a fractogram from which cell numbers and their size or mass distribution can be calculated. A direct method for estimating bacterial biomass (dry organic matter content) which offers distinct advantages over present methods for calculating biomass has been developed.