CD40L/IL-4 activated clonal isotype expression in healthy donor, WM, and IgM MGUS cells. Clonal isotype transcripts were determined by RT-PCR using standard (Std) or enhanced (Enh) amplification protocol (A). Day 0, unstimulated; day 6, after stimulation; CDR2/CDR3 (lane 1), CDR2/Cμ (lane 2), CDR2/Cδ (lane 3), CDR2/Cα (lane 4), and CDR2/Cγ (lane 5). In normal control, lanes 1 and 3 are not determined and CDR2 is replaced by FR1c. In WM1-09, CDR2 is replaced by CDR3R. Representative results of germline transcript up-regulation after CD40L/IL-4 activation are shown in panel B. The primers (sense/antisense) and expected size of PCR products are as follows: M, sIμ/asCμ, 537 bp; G1, sIγ1/2/asCγ1, 603 bp; G2, sIγ1/2/asCγ2, 597 bp; G3, sIγ3/asCγ3, 670 bp; G4, sIγ4/asCγ4, 411 bp; E, sIϵ/asCϵ, 125 bp. CDR3 repertoire analysis of WM1-17 was conducted for B cells expressing each isotype (FR3/Cμ, FR3/Cδ, FR3/Cα, and FR3/Cγ) and total B cells (FR3/JHc) (C). IgM (CDR2/Cμ), IgG (CDR2/Cγ), and total clonal frequency (CDR2/CDR3) was analyzed in poststimulated CD20⁺ cells of WM1-17 (D).