CD38⁺⁺CD20^± B cells express genes indicative of commitment to the plasma cell lineage and derivation from germinal centers. (A-E) RNA was extracted from sort-purified naive (CD20⁺CD27^–), memory (CD20⁺CD27⁺), and CD38⁺⁺CD20^± (ISC) B cells and transcribed into cDNA. Amounts of cDNA from the different populations were normalized for expression of GAPDH (E) and then used as a template to determine the relative expression levels of (A) Blimp-1, (B) XBP-1, (C) BSAP, and (D) Bcl-6 by sqPCR. Molecular grade dH₂O was used as a negative control. (F) Immunoglobulin V_H5 genes were amplified from CD38⁺⁺CD20^± B cell cDNA, cloned, and sequenced. Each line represents a single V_H5 gene. Sequences with the no. 1 and no. 2 prefixes were derived from CD38⁺⁺CD20^± B cells sort-purified from 2 separate donor spleens. Vertical bars represent silent mutations; vertical bars with • represent replacement mutations. The total number of mutations detected in the different cloned genes is shown at the end of each sequence. The mutation frequency, percentage replacement mutations and R/S ratio within the entire immunoglobulin V_H5 gene sequence and individual FR and CDR are indicated. These values were calculated for immunoglobulin V_H5 sequences that contained somatic mutations. *Significant increase (P < .001) in the frequency of mutation in CDR1 compared with other regions and the total V_H5 gene.