The Effect of Washing After Protein Blocking on Immunohistochemical Staining Applied to Paraffin Sections

Authors : Mete Keçeci, Elif Koca, Esra Arslan, Reyyan Başak Denek, Ezgi Su Sari, Ipek Pınar Akpinar, Bünyamin Ülker, Kağan Atici, Özgenur Erbakan, Osman Cengil
Pages : 325-330
Doi:10.29058/mjwbs.1337146
View : 64 | Download : 67
Publication Date : 2023-12-31
Article Type : Research Paper
Abstract :Aim: Immunohistochemistry is still frequently preferred in both diagnostic and experimental studies because it can show proteins where they are in the tissue. One of the main problems in immunohistochemistry is the background staining, which can be prevented by protein blocking and which occurs as a result of binding of primary antibodies to tissue proteins and Fc receptors in the tissue due to antigenic similarity. There is no consensus on whether to wash after protein blocking in current publications and immunohistochemistry manuals published by manufacturers. Material and Methods: In our study, routine immunohistochemistry procedure was applied to determine the expression of TNF-α on 5 μm thick sections obtained from rat gastric tissue samples in which an experimental gastric ulcer model was created with ethanol, and two groups were formed, with and without washing after protein blocking, with 10 sections in each group. For semi-quantitative evaluation, the histological score (h-score) was calculated from the images obtained from the immunohistochemically stained preparations of both groups and the obtained data were statistically compared. Results: As a result of our study, no statistically significant difference was found between the h-score values of the groups that were washed after protein blocking in immunohistochemical staining and the groups that did not wash (p=0.971). The median (min-max) values of the groups with and without washing are 211 (179-244) and 215 (171-251), respectively. Conclusion: In the immunohistochemical staining procedure, washing after protein blocking does not change the staining intensity, does not create background staining, and does not affect the result of the h-score for semiquantitative evaluation. Similar studies are recommended for other species, tissue types and antigens.
Keywords : İmmünohistokimya, Parafin kesit, Protein bloklama, Yıkama, h skor

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