Solubility of cytoskeletal proteins in immunohistochemistry and the influence of fixation.

Details

Serval ID
serval:BIB_1E04A2803E44
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Solubility of cytoskeletal proteins in immunohistochemistry and the influence of fixation.
Journal
Journal of Histochemistry and Cytochemistry
Author(s)
Riederer B.M., Porchet R., Marugg R.A., Binder L.I.
ISSN
0022-1554 (Print)
ISSN-L
0022-1554
Publication state
Published
Issued date
1993
Volume
41
Number
4
Pages
609-616
Language
english
Abstract
For accurate and quantitative immunohistochemical localization of antigens it is crucial to know the solubility of tissue proteins and their degree of loss during processing. In this study we focused on the solubility of several cytoskeletal proteins in cat brain tissue at various ages and their loss during immunohistochemical procedures. We further examined whether fixation affected either solubility or immunocytochemical detectability of several cytoskeletal proteins. An assay was designed to measure the solubility of cytoskeletal proteins in cryostat sections. Quantity and quality of proteins lost or remaining in tissue were measured and analyzed by electrophoresis and immunoblots. Most microtubule proteins were found to be soluble in unfixed and alcohol fixed tissues. Furthermore, the microtubule proteins remaining in the tissue had a changed cellular distribution. In contrast, brain spectrin and all three neurofilament subunits were insoluble and remained in the tissue, allowing their immunocytochemical localization in alcohol-fixed tissue. Synapsin I, a protein associated with the spectrin cytoskeleton, was soluble, and aldehyde fixation is advised for its immunohistochemical localization. With aldehyde fixation, the immunoreactivity of some antibodies against neurofilament proteins was reduced in axons unveiling novel immunogenic sites in nuclei that may represent artifacts of fixation. In conclusion, protein solubility and the effects of fixation are influential factors in cytoskeletal immunohistochemistry, and should be considered before assessments for a quantitative distribution are made.
Keywords
Animals, Cats, Cytoskeletal Proteins/analysis, Cytoskeletal Proteins/chemistry, Formaldehyde, Frozen Sections, Immunohistochemistry, Microtubule-Associated Proteins/analysis, Microtubule-Associated Proteins/chemistry, Polymers, Solubility, Tissue Fixation, Tubulin/analysis, Tubulin/chemistry, Visual Cortex/chemistry
Pubmed
Web of science
Create date
24/01/2008 14:34
Last modification date
20/08/2019 12:54
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