Confocal scanning light microscopy of the Escherichia coli nucleoid: comparison with phase-contrast and electron microscope images

Abstract

The nucleoid of living and OsO4- or glutaraldehyde-fixed cells of E. coli strains was studied with a phase-contrast microscopy, confocal scanning light microscopy and EM. The trustworthiness of the images obtained with the confocal scanning light microscope was investigated by comparison with phase-contrast micrographs and reconstructions based on serially sectioned material of DNA-containing and DNA-less cells. This comparison showed higher resolution of the confocal scanning light microscope as compared with the phase-contrast microscope and agreement with results obtained with EM. The effects of fixation on the structure of the nucleoid were studied in E. coli B/r H266. Confocal scanning light micrographs and EM reconstructions showed that the shape of the nucleoid remained similar after OsO4 or glutaraldehyde fixation; however; the OsO4 nucleoid appeared to be somewhat smaller and more centralized within the cell.