Visualization of vaginal flora in cervical smears using a modified microwave silver-staining method.

The advantage of studying the vaginal flora to determine the bacteria and fungi present in cervical smears (as opposed to cultivation of these micro-organisms) is that the micro-organisms can be observed in their natural habitat. However, they are only faintly stained by the conventional Papanicolaou method. Accordingly, contrast is weak and visualization poor. For this reason, we developed a modified microwave silver-staining method that can be performed retrospectively on stained smears. Bacteria and fungi stain distinctly black and can be studied in greater detail, and their inter-relationship can be visualized. Haematoxylin or Eosin counterstain allows us to visualize vaginal inhabitants in relation to epithelial cells. In the series presented here, we show that a modified microwave silver-staining method is well suited to studying the ecology of micro-organisms in smears taken from women presenting to their doctor with clinical symptoms. Using this staining method, we have shown that lactobacilli overgrowth is associated with symptoms.

Sporicidal activity of chemical and physical tissue fixation methods.

AIMS: The effects of alcohol based fixation and microwave stimulated alcohol fixation were investigated on spores of Bacillus stearothermophilus and Bacillus subtilis (var. niger). METHODS: Spores were exposed to 10% formalin, or different concentrations of various alcohol containing fixatives (Kryofix/Spuitfix). Adequate controls were also set up in conjunction with the test solutions. The spores were immersed with and without adjunctive microwave stimulation in the various solutions tested. Possible surviving spores were recovered in revival broth and after incubation, and Gram staining viable counts were performed. RESULTS: Alcohol based fixatives did not have a sporicidal effect on B stearothermophilus or B subtilis (var. niger) spores, and microwave stimulated alcohol fixation at 450 W and up to 75 degrees C did not have a sporicidal effect. CONCLUSIONS: When alcohol based fixatives are used for fixation, precautions should be taken with the material thus treated, as it may contain viable spores or other pathogens, which are destroyed after 24 hours of formalin treatment. Of the physicochemical methods tested involving microwaving, none was successful in eliminating viable spores from the test material.

Upgrading unsatisfactory cervical smears with the MiB-1 method.

When an unsatisfactory smear (thick inflammatory infiltrate or blood influencing the staining characteristics of the epithelial cells) is restained for MiB-1, the diagnostic proliferating cells are visualized, and the MiB-1-positive smears can be thus upgraded as borderline, grade I, II, and III, corresponding with the cytologic diagnoses of, respectively, ASCUS, CIN I, II, and > or = III. In a period of 18 months, 2,068 unsatisfactory smears out of a material of 84,817 smears were restained for MiB-1. In the unsatisfactory group, significantly more abnormal smears were detected than in the satisfactory group. Seventy-five of the unsatisfactory group were biopsied because of the MiB-1 findings: Three women proved to have severe dysplasia, four had carcinoma in situ, and three had invasive carcinoma. The per mileage for invasive cervical carcinoma was ten times larger in the unsatisfactory group than in the satisfactory group; thus the MiB-1 method has further enhanced our diagnostic acumen in this difficult type of smears.