Effects of treatment on differential staining of BrdU labeled metaphase chromosomes: three-way differentiation of M3 chromosomes

Abstract

In this paper methodology is described which yields three-way Giemsa 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 third) 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. Using these methods it has been demonstrated that approximately twice as many SCEs occur during the first S-period in the presence of the drug (labeling=B1T0XT0B1)1 as occur during the second S-period (labeling=B2B1XT0B2)1. The three-way differentiation pattern is thought to result from a stepwise decrease in the amount of BrdU incorporated during the first, second and third 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).

Presence of abnormally high incidences of sister chromatid exchanges in three successive cell cycles in Bloom's syndrome lymphocytes

“…In three-way differentially stained M3 cells, half regions of chromosomal complements consisted of darkly stained and intermediately stained chromatids and the other half consisted of intermediately stained and lightly stained chromatids. Using these metaphases, SCEs occurring during $1, $2 and $3 could be distinguished from one another and scored (Miller 1976;Schvartzman 1979;Tsuji and Kato 1981).…”

“…In the present study the SCEs occurring during each cell cycle were counted separately in three-way differentially stained, thirdcycle (M3) metaphases (Miller et al 1976;Schvartzman 1979); Tsuji and Kato 1981). Another method, two-way differentiation of M3 metaphases, allows SCEs occurring during the first two cell cycles to be distinguished from the SCEs occurring during the third cycle (Geard 1974;Tice et al 1975;Wolff and Perry 1975).…”

Chromium-Induced Sister Chromatid Exchanges in CHO Cells

“…If SCEs were induced by BUdR (MILLER et al 1976;SCHWARTZMAN et al 1979 ), the expected ratio of asymmetrical to symmetrical SCEs would be 3,3: 1 (ScHWARTZMAN et al 1979), since their frequency would vary with the number of replication periods that cells developed in the presence of BUdR.…”

Aphidicolin-Resistant Mutants of Mouse Lymphoma L5178y Cells With a High Incidence of Spontaneous Sister Chromatid Exchanges

“…With these methods, almost all of the metaphases exhibited three-way differentiation of chromosomes, but not a replication kinetics pattern. SCEs in each cell cycle (Sl-, S2-and S3-SCEs) were all reciprocal types of exchanges and could be counted separately based on the patterns of three-way differentiation (MILLER, ARONSON and NICHOLS 1976;SCHVARTZMAN 1979;SCHVARTZMAN and GOYANE~ 1980;TSUJI and KATO 1981).…”