Short μ transcripts are present in L^−/− plasma cells. (A) Spleen cells from a 6-month-old L^−/− and a NM mouse were stained either for B220 and IgM or B220 and CD138, and sorted using the indicated gates. IgM⁺ cells from L^−/− spleen represent 0.2% of cells in the lymphocyte gate. (B-C) RT-PCR analysis of the sorted fractions using a degenerate V_H (Vgen) or J_H (JH3) primer,²¹  in combination with a Cμ3 or Cμ2 reverse primer, respectively. H₂O served as negative control and the white line indicates a repositioned marker lane from the same gel. RNA was prepared and reverse transcribed from the sorted cell fractions (NM and L^−/−) as indicated (B220⁺, B220 int[ermediate], B220 high, μ⁺, and CD138⁺) and from total spleen aliquots as control. The cDNA equivalent of 200 cells was used in each assay, except for the μ⁺ fraction (100 cells) and the L^−/− CD138⁺ fraction (25 cells). (B) A normal μ transcript results in a ∼ 1.1-kb fragment after V_H-Cμ3 amplification. The small band of ∼ 0.45 kb, found only in L^−/− mice, corresponds to a transcript without C_H1 and C_H2, and the intermediate band ∼ 0.8 kb corresponds to a transcript lacking C_H1. (C) After J_H-Cμ2 amplification, a normal band of ∼ 0.7 kb is found, which is reduced to ∼ 0.35 kb, lacking C_H1, in L^−/− total spleen or CD138⁺ cells. A weak band is also seen in μ⁺ cells.