In vitro BrdUrd incorporation of colorectal tumour tissue

This study describes a novel in vitro method for the incorporation of the thymidine analogue, bromodeoxyuridine (BrdUrd), in fresh colorectal tumour tissue. Disaggregation by pronase, collagenase and DNAse resulted in high cell yields of viable single cell suspensions, representative of the original tumour, which could be infiltrated with BrdUrd. A modified ELISA identified optimal incubation times and BrdUrd concentrations. This technique has been used in preliminary studies to investigate two important areas intrinsic in the analysis of BrdUrd colorectal cell proliferation data: 1) to determine the effects of the individual constituents of the cell culture media, in particular glutamine, on BrdUrd incorporation in suspensions of colorectal cells and 2) to examine the denaturation step. This method will have wide applicability in investigations of cell proliferation status in both normal and diseased tissue.