期刊
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY
卷 58, 期 1, 页码 29-39出版社
SAGE PUBLICATIONS LTD
DOI: 10.1369/jhc.2009.954354
关键词
extraction; fixation; in situ zymography; matrix metalloproteases; morphology
类别
资金
- Norwegian Cancer Society
- Northern Norwegian Regional Health Authorities
- Erna and Olav Aakre Foundation for Cancer Research
In situ zymography is a method for the detection and localization of enzymatic activity in tissue sections. This method is used with frozen sections because routine fixation of tissue in neutral-buffered formalin inhibits enzyme activity. However, frozen sections present with poor tissue morphology, making precise localization of enzymatic activity difficult to determine. Ethanol- and zinc-buffered fixative (ZBF) are known to preserve both morphological and functional properties of the tissue well, but it has not previously been shown that these fixatives preserve enzyme activity. In the present study, we show that in situ zymography can be performed on ethanol- and ZBF-fixed paraffin-embedded tissue. Compared with snap-frozen tissue, ethanol- and ZBF-fixed tissue showed stronger signals and superior morphology, allowing for a much more precise detection of gelatinolytic activity. Gelatinolytic enzymes could also be extracted from both ethanol- and ZBF-fixed tissue. The yield, as analyzed by SDS-PAGE gelatin zymography and Western blotting, was influenced by the composition of the extraction buffer, but was generally lower than that obtained from unfixed tissue. (J Histochem Cytochem 58:29-39, 2010)
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