EFFECTS OF TREATMENT ON DIFFERENTIAL STAINING OF BRDU LABELED METAPHASE CHROMOSOMES - 3-WAY DIFFERENTIATION OF M3 CHROMOSOMES

Methodology is described which yields 3-way Giesma differentiation (light-medium-dark) in human metaphase chromosomes exposed to 5-bromodeoxyuridine (BrdU) for 3 DNA synthetic periods (or exposed for 2 DNA synthetic periods and removed from exposure for the 3rd) by means of which all of the sister chromatid exchanges (SCEs) occurring during (or shortly after) S1, S2 and S3 can be accurately counted and distinguished from one another. Approximately twice as many SCEs occur during the 1st S-period in the presence of the drug (labeling = B1T0 .times. T0B1)1 as occur during the 2nd S-period (labeling = B2B1 .times. T0B2)1. THe 3-way differentiation pattern is thought to result from a stepwise decrease in the amount of BrdU incorporated during the 1st, 2nd and 3rd DNA synthetic periods. These methods can also be used to differentiate between unlabeled (T2T0) and unifilarly labeled (B1T2) sister chromatids and are potentially useful in the detection of sub-chromatid exchanges (none were detected).