Immunocytochemical detection of 5-bromo-2-deoxyuridine incorporation in individual cells

5-Bromo-2-deoxyuridine (BrdUrd) is a thymidine analogue which can be used to detect DNA synthesis in cells. The present paper describes a method for the fixation and DNA denaturation of cells that affects neither the morphology of the cells not their adhesion to a glass surface. The cells were fixed in absolute ethanol and glacial acetic acid followed by gradual rehydration. Denaturation of DNA was achieved by very brief exposure of the cells to low pH, followed by heating to high temperature and rapid cooling. Incorporation of BrdUrd was assessed from the binding of monoclonal antibody to BrdUrd in single-stranded DNA as detected with an immunocytochemical method. This method makes it possible to demonstrate BrdUrd labeling of individual, morphologically identifiable cells. The results obtained with this method correspond well with the autoradiographic data on [3H]thymidine incorporation.