LRCs can be serially passaged and can generate MM plasma cells. (A) Mononuclear cells were isolated from the mobilized blood preparations, labeled with CFSE, grown in 3-D for 14 days, after which CFSE^high lymphocytic cells were sorted from the harvested cells, and cultured in a CFU assay (no morphologic differences between clonotypic and nonclonotypic colonies were observed). Clonotypic or nonclonotypic progeny were generated from LRCs in CFU assay and tested for clonotypic IgH VDJ signatures by PCR and by RQ-PCR. Representative single-stage PCR analysis of the clonotypic IgH VDJ and β2-microglobulin from representative CFU progeny is shown; these results were confirmed using RQ-PCR (Figure 6D and data not shown). (B) LRCs were sorted and placed into CFU culture, followed by serial replating for 6 generations. Each panel is a representative image from the consecutive passage of a colony (p1) from a single patient (n = 6 patients). (C) FACS analysis of the LRC progeny after the sixth serial passage of a clonotypic colony (culture p6 from panel B). The entirety of p4 and p5 was used for clonal expansion to obtain sufficient cells for the p6 FACS analysis. Histograms are representative of those obtained for 6 different patients. All of the secondary colonies from clonotypic CFU were clonotypic. In contrast, secondary colonies from the nonclonotypic CFU were all nonclonotypic. Self-renewal capability within colonies was further supported by the morphology of the CFU arising from LRCs, which at all passages included small secondary colonies scattered throughout the primary colony, as shown in the figure.