Enrichment of colony forming cells by methotrexate and isokinetic density gradient centrifugation

Purification of target cells responsive to granulopoietic regulators would be of importance in understanding their mechanism of action. Previous studies have shown that mouse marrow could be enriched 3- to 7-fold with respect to agar colony forming cells (CFU-C) by appropriately scheduled doses of Methotrexate. To further increase this enrichment, such marrow has been subjected to centrifugation through an isokinetic gradient of Ficoll in culture medium. Under optimal conditions (17.5 min at 77.5 X g) the combination of chemotherapy and velocity sedimentation yielded a 17-fold enrichment of CFU-C's.