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. 1978 Apr;75(4):1820–1824. doi: 10.1073/pnas.75.4.1820

Cytoplasmic microtubular images in glutaraldehyde-fixed tissue culture cells by electron microscopy and by immunofluorescence microscopy

Klaus Weber 1, Peter C Rathke 1, Mary Osborn 1
PMCID: PMC392432  PMID: 417343

Abstract

Electron microscopy and indirect immunofluorescence microscopy using monospecific tubulin antibodies were performed in parallel on glutaraldehyde-fixed tissue culture cells without osmium fixation. In order to reduce the excess aldehyde groups of the strongly crosslinked cellular matrix, which normally interfere with subsequent immunofluorescence microscopy, a mild NaBH4 treatment was introduced during or after the dehydration steps. Cells processed through the NaBH4 step show, in transmission electron microscopy, normal cytoplasmic microtubules approximately 250 Å in diameter. When such cells are subjected to indirect immunofluorescence microscopy using monospecific tubulin antibody they reveal a complex system of unbroken, fine, fluorescent fibers traversing the cytoplasm between the perinuclear space and the plasma membrane. Thin sections of cells processed through the indirect immunofluorescence procedure show antibody-decorated microtubules with a diameter of approximately 600 Å. This decoration is not obtained when non-immune IgGs are used instead of monospecific antitubulin IgGs. Thus, a direct comparison of cytoplasmic microtubules in glutaraldehyde-fixed cells by both electron microscopy and immunofluorescence microscopy can be obtained.

Keywords: tubulin antibody, NaBH4, fixation procedures, actin, immunocytochemistry

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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