Comparison of Antigen Retrieval Techniques in Immunohistochemistry

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Em-orn Phanomsri
Nisana Namwat
Sakda Waraasawapati
Roongpet Tangrassameeprasert
Supinda Koonmee


Objective: The study aimed to find the best antigen retrieval buffer and heat source for the immunohistochemical staining of various kind of antibodies in our laboratory.

Materials and methods: We designed the method to evaluate the efficacy of three different retrieval solutions in cluding 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. Morever, 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



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Phanomsri E- orn, Namwat N, Waraasawapati S, Tangrassameeprasert R, Koonmee S. Comparison of Antigen Retrieval Techniques in Immunohistochemistry. SRIMEDJ [Internet]. 2013 Nov. 1 [cited 2022 Dec. 10];24(2):130-6. Available from:
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