A plasma clot culture system for growing and antiproliferative drug sensitivity testing of myeloma stem cells.

An easy-to-perform clonogenic culture system for myeloma stem cells is presented which approaches the patient's in vivo situation more closely and therefore seems particularly well-suited for antiproliferative drug sensitivity assays. Bone marrow cells are propagated in clotted autologous or allogenic plasma that is enriched with 2-mercaptoethanol, insulin, and synthetic nucleotides and co-factors for nuclear synthesis, no feeder layer or conditioned medium is necessary. Clusters and colonies consisting of between 8 and 60 cells readily formed within 6-8 days after cloning yielding a plating efficiency between 0.12 and 2.16%. A linear relationship between the number of cells plated and colony formation was found from 10(5) through 2 X 10(6) cells plated. The successful growth rate for 65 tests from 53 patients amounted to 90.8%. Morphological and histochemical examination of the clusters revealed lymphoid cells at various stages of maturation ranging from lymphocytic and lymphoblastoid to lymphoplasmacytic and plasma cells. Tumor origin of the clones was demonstrated by immunofluorescence studies in which Ig-positive cells stained only for the heavy and light chain isotypes identical to those of the patient's serum paraprotein. Anti-idiotypic antisera confirmed the patient's specific malignant phenotype of the colonies formed. The technique of the assay is described in detail.