Webmedcentral - Immunohistochemistry ArticlesThe Immunohistochemistry articles published by Webmedcentral
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2024-03-29T07:08:22+01:00webmedcentral logo
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http://www.webmedcentral.com/images/Header_Logo.giftext/html2011-01-07T12:11:05+01:00http://www.webmedcentral.com/Prof. Jeanne Adiwinata PawitanImportance Of Secondary Antibody Choice In Immunohistochemistry: Learning From A Failure
http://www.webmedcentral.com/article_view/1453
Aim: to optimize Tenascin-C immunohistochemistry staining protocol for rat cardiac tissueMethods: This is a preliminary experimental study at Histology Lab Faculty of Medicine University of Indonesia from 7th December 2007 to 2nd July 2008. Samples were paraffin sections of rat cardiac tissues, and paraffin sections of brain tissue were used as positive control and absence of overnight primary antibody incubation as negative control. Formalin fixation was used before paraffin blocks were made. Immunohisto-chemistry protocol was modified from Hannekamp, et al, who stained human tissue.Staining results were recorded and analyzed. Staining was considered successful when the positive and negative controls were positive and negative respectively. If the protocol was considered unsuccessful, the data obtained and the steps in the protocol was reviewed and analyzed for cause/s of failure and troubleshooting steps were taken.Results: The staining results according to the protocol and subsequent troubleshooting protocols showed mostly false positive results, which cause was supposed to be the cross reaction of secondary antibody with the rat tissue.Conclusion: In starting a new immunohistochemistry staining for paraffin sections, care should be paid on the compatibility of the kit for paraffin section, choice of primary antibody that react with the antigen to be studied, and choice of secondary antibody that do not react with the tissue to be studied.