Comparison of Antigen Retrieval Techniques in Immunohistochemistry

ABSTRACT

Objective: The study aimed to find the best antigen retrieval buffer and heat source for the immunohistochemical staining of various kinds of antibodies in our laboratory.Materials and methods: We designed the method to evaluate the efficacy of three different retrieval solutions including 10mM Tris-HCl + 1mM EDTA, pH 9, 0.05%citraconic anhydride solution pH 7.4 and 10 mM citrate buffer pH 6, and 3 heat source pretreatment methods (Microwave, pressure cooker and water bath treatment)to retrieve twenty-one immunoreactivity in formalin-fixed, paraffin-embedded sections.Results: We found that, modified retrieval solution, 10mM Tris-HCl + 1mM EDTA buffer, pH 9 is the most efficient for a large variety of antibodies and not depending on heat sources. On the other hand, 0.05% citraconic anhydride solution and 10mM citrate buffer are moderate and poor retrieval solutions, respectively. Moreover, these two solutions are heat source-dependent.Conclusion: These results demonstrate that 10mM Tris-HCl + 1mM EDTA, pH 9 and heat-pretreatment is useful for the immunohistochemistry of many antigens in aldehyde-fixed, paraffin-embedded tissues. Keywords antigen retrieval, immunohistochemistry, citraconic anhydride, formaldehyde