Methotrexate-leucovorin synchronization of human bone marrow cultures. Induction of high-resolution R- and G-banding.

Simple R- and G-banding techniques based on a methotrexate S-phase block, released by exposure to leucovorin, were used for preparation of human bone marrow samples. R-bands were induced by adding 5-bromodeoxyuridine and Hoechst 33258 to the cultures, followed by a modified FPG-staining. G-bands were induced by adding ethidium bromide to the cultures, followed by a modified trypsin treatment of the slides. Due to Hoechst 33258 or ethidium bromide exposure prior to harvest chromosome contraction was partly inhibited and well-defined R- or G-banding was observed after appropriate staining in elongated prometaphase chromosomes.