Unstained and in vivo fluorescently stained bacterial nucleoids and plasmolysis observed by a new specimen preparation method for high-power light microscopy of metabolically active cells.
Microscope slides were coated with a layer of gelatin, the thickness of the gelatin increasing linearly along the long axis. The bacterial suspension is applied to the dried gelatin and covered by a coverslip. The medium is absorbed by the gelatin and thus the cells applied against the coverslip. By this method, cultures of concentrations below 10(8) cells/ml provide statistically relevant numbers for observation without prior concentration steps. It is easier to apply than the existing methods for the observation of bacterial nucleoids by phase contrast imaging. Because the cells are maintained in growing conditions the method is useful for the vital fluorescence DAPI-staining of various bacterial species and for observations of plasmolysis and its reversal at different physiological conditions and extracellular osmolalities. The previously generally assumed view that the plasmolytic changes of the cell morphology are immediate upon the hyperosmotic shock and are rapidly repaired when the cell is able to metabolize actively was confirmed; this is in contrast to some recent claims.
Kellenberger
E
E
Institut de Génétique et de Biologie Microbiennes, Université de Lausanne, Switzerland.
Kellenberger-Van der Kamp
C
C
D001407:Bacillus / Q000166:cytology*; D001433:Bacteriolysis; D004926:Escherichia coli / Q000166:cytology* / Q000648:ultrastructure; D005456:Fluorescent Dyes; D005780:Gelatin; D007211:Indoles; D008856:Microscopy, Fluorescence / Q000379:methods*; D008858:Microscopy, Phase-Contrast; D009997:Osmotic Pressure
D005456:Fluorescent Dyes; D007211:Indoles; C007293:DAPI; D005780:Gelatin