Generalized blocking in S phase by methotrexate.

An attempt was made to enhance the frequency of prometaphase cells for high-resolution-banding studies in untransformed Syrian hamster fibroblasts using a typical methotrexate (MTX) block/bromodeoxyuridine release schedule. The recovery 'wave' was serially sampled and detailed sub-phase analysis made using the replication bands resulting from bromodeoxyuridine uptake. Of the 3 batches of MTX used, one (Sigma greater than 2 years old) was found to have decayed to a non-toxic compound which produced almost no measurable perturbation of the cell cycle at any concentration used. The other 2 (new Sigma and new Lederle), whilst producing mitotic index fluctuations which could be construed to indicate blocking and "synchrony", gave absolutely no evidence of any specific blocking site, but rather a general stoppage (or slowing down) during MTX treatment and continuation in exactly the same order as untreated controls upon release.