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Blood, Vol. 104, Issue 12, 3647-3654, December 1, 2004
Human blood IgM "memory" B cells are circulating splenic marginal zone B cells harboring a prediversified immunoglobulin repertoire
Blood Weller et al.
104: 3647
Supplemental materials for: Weller et al, Vol 104, Issue 12, 3647-3654
Files in this Data Supplement:
- Figure 1. CDR3 spectratyping of V3-15-Cµ transcripts (JPG, 68 KB)
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CDR3 size distribution analysis (spectratyping) of V3-15-Cµ transcripts expressed in peripheral and splenic IgM+IgD+CD27+ B cells of a 9-year-old child before and after immunization with a polysaccharidic vaccine. Polymerase chain reaction (PCR) amplification of the CDR3 region of V3-15-Cµ transcripts was achieved by a 2-step strategy. First, cDNA from sorted IgM+IgD+CD27+ B cells was amplified using a sense V3-15-leader-specific primer with an antisense primer specific for the constant region Cµ The resulting PCR products were then amplified using a sense V3-15-FR3 specific primer with the same antisense Cµ primer. After electrophoresis on a 6% denaturing polyacrylamide gel, PCR products were revealed by silver staining. The bands indicated by arrows corresponding to one particular CDR3 length were excised, reamplified, cloned, and sequenced.
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