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Cell proliferation assay – method optimisation for in vivo labeling of DNA in the rat forestomach

Joksić, Gordana, Mićić, Mileva, Filipović, Jelena, Drakulić, Dunja, Stanojlović, Miloš, Čalija, Bojan, Valenta Šobot, Ana, Demajo, Miroslav, Nilsson, Robert
Acta veterinaria 2017 v.67 no.1 pp. 1-10
DNA, adults, cell proliferation, forestomach, immunohistochemistry, pathophysiology, pharmacology, rats, toxicology
The study of cell proliferation is a useful tool in the fields of toxicology, pathophysiology and pharmacology. Cell proliferation and its degree can be evaluated using 5-bromo-2′-deoxyuridine which is incorporated into the newly synthesized DNA. The aim of this study was the optimization of subcutaneous application of 5-bromo-2′-deoxyuridine implantation for continuous and persistent marking of proliferating cells in the rat forestomach. 3-tert-Butyl-4-hydroxyanisole was used as the agent that ensures cell proliferation. In order to determine the optimal dose for proliferating cells labeling, 5-bromo-2′-deoxyuridine doses of 50 mg, 100 mg, 200 mg or 350 mg were implemented 2 days prior to sacrifice by flat-faced cylindrical matrices. Immunohistochemical analysis using 5-bromo-2′-deoxyuridine in situ detection kit was performed for the detection of 5-bromo-2′-deoxyuridine labeled cells. The results showed that for adult rats, the optimum 5-bromo-2′-deoxyuridine dose is 200 mg per animal for subcutaneous application. The here described manner of 5-bromo-2′-deoxyuridine in vivo labeling provides a simple, efficient, and reliable method for cell labeling, and at the same minimizes stress to animals.