HYDROCHLORIC-ACID DENATURATION OF COLORECTAL TUMOR-TISSUE INFILTRATEDWITH BROMODEOXYURIDINE

The thymidine analogue, bromodeoxyuridine, is substituted into DNA dur ing DNA synthesis and can be used to identify those cells which have p assed through the S-phase of the cell cycle. Incorporated bromodeoxyur idine can be detected using anti-bromodeoxyuridine monoclonal antibodi es which bind to the exposed bromodeoxyuridine in single-stranded DNA after a suitable denaturation step. Hydrochloric acid is the most comm only employed denaturation agent in bromodeoxyuridine monoclonal antib ody methodologies. This preliminary study was to validate the hydrochl oric acid denaturation step for colorectal tumour tissue infiltrated i n vivo with bromodeoxyuridine. Standard immunohistochemistry and flow cytometric techniques using Bu20a were employed across a range of hydr ochloric acid concentrations. Although high labelling indices were ach ieved using acid concentrations of 0.10 M HCl, the optimal hydrochlori c acid concentration was not the same in all tumours. Sensitivity of D NA to hydrochloric acid denaturation should be carefully considered in bromodeoxyuridine methodologies using the monoclonal antibody Bu20a. (C) 1994 Wiley-Liss, Inc.