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Optimization of fixation methods for observation of bacterial cell morphology and surface ultrastructures by atomic force microscopy.

ABSTRACT: Fixation ability of five common fixation solutions, including 2.5% glutaraldehyde, 10% formalin, 4% paraformaldehyde, methanol/acetone (1:1), and ethanol/acetic acid (3:1) were evaluated by using atomic force microscopy in the present study. Three model bacteria, i.e., Escherichia coli, Pseudomonas putida, and Bacillus subtilis were applied to observe the above fixation methods for the morphology preservation of bacterial cells and surface ultrastructures. All the fixation methods could effectively preserve cell morphology. However, for preserving bacterial surface ultrastructures, the methods applying aldehyde fixations performed much better than those using alcohols, since the alcohols could detach the surface filaments (i.e., flagella and pili) significantly. Based on the quantitative and qualitative assessments, the 2.5% glutaraldehyde was proposed as a promising fixation solution both for observing morphology of both bacterial cell and surface ultrastructures, while the methonal/acetone mixture was the worst fixation solution which may obtain unreliable results.

SUBMITTER: Chao Y

PROVIDER: S-EPMC3181414 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Optimization of fixation methods for observation of bacterial cell morphology and surface ultrastructures by atomic force microscopy.

Chao Yuanqing Y Zhang Tong T

Applied microbiology and biotechnology 20110901 2

Fixation ability of five common fixation solutions, including 2.5% glutaraldehyde, 10% formalin, 4% paraformaldehyde, methanol/acetone (1:1), and ethanol/acetic acid (3:1) were evaluated by using atomic force microscopy in the present study. Three model bacteria, i.e., Escherichia coli, Pseudomonas putida, and Bacillus subtilis were applied to observe the above fixation methods for the morphology preservation of bacterial cells and surface ultrastructures. All the fixation methods could effectiv ...[more]

PMID: 21881891

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Project description:BackgroundLichen planopilaris (LPP) is a chronic lymphocytic skin disease manifested by progressive scarring alopecia. The diagnosis of LPP is made based on histopathological examination, although it is not always definite. The current study evaluates the effectiveness of non-invasive atomic force microscopy (AFM) hair examination in detecting morphological differences between healthy and diseased hair.Materials and methodsHere, three to five hairs from lesional skin of 10 LPP patients were collected and examined at nine locations using AFM. At least four images were taken at each of the nine sites. Metric measurements were taken and metric (length, width, and scale step height) and morphological features (striated and smooth surface of scales, the presence of endocuticle and cortex, shape of scales edges, scratches, pitting, cracks, globules, and wavy edge) were compared with hair from healthy controls. In addition, areas on diseased hair where the process of pathological, unnatural delamination of the hair fiber occurs are described.ResultsThere was a statistically significant difference in the number of scratches in the initial sections of the LPP hair, in the intensity of wavy edges along the entire length of the tested hair, and in the number of scales with pitting in the middle section of the hair. In addition, a statistically significant higher number of scales with striated surface was found in LPP group starting at 3.5 cm from the root continuing towards the free end of the hair. Other morphological changes such as presence of cortex, globules, oval indentations, and rod-like macrofibrillar elements were also assessed, however, detailed results are not presented, as the differences shown in the number of these morphological changes were not significantly different.ConclusionThis publication outlines the differences between virgin, healthy Caucasian hair, and the hair of LPP patients. The results of this study can be used for further research and work related to LPP. This is the first attempt to characterize the hair of LPP patients using AFM.

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Optimization of fixation methods for observation of bacterial cell morphology and surface ultrastructures by atomic force microscopy.

Publications

Optimization of fixation methods for observation of bacterial cell morphology and surface ultrastructures by atomic force microscopy.

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