Summary of Single-Cell Analysis of Two Cases for V Gene Rearrangements
Patient No. . | Experiment No. . | Cells Positive for V Rearrangements After PCR With Different Forward Primers . | PCR Products . | Rearrangements . | |||||
---|---|---|---|---|---|---|---|---|---|
Vκ . | VHFRI . | VHFRII . | Clone- Specific . | Total . | Sequenced . | Repeated . | Unique . | ||
1 | 1a | ND | 13/17 | —-150 | ND | 13 | 13 | 13 VH1 | |
1b | ND | 7/14 | —-150 | 7/14 | 7 | 7 | 7 VH1 | ||
2 | 2′LN | 0/13 | 0/13 | —-150 | 3/13 | 3 | 3 | 3 VH1 | |
2"LN | 0/7 | —-150 | 1/7 | 3/7 | 3 | 3 | 3 VH1 | ||
2"spleen | 0/10 | —-150 | 1/10 | 2/10 | 2 | 2 | 2 VH1 | ||
Control | 1a-151 | ND | 6/8 T cells | —-150 | ND | 8 | 8 | 5 VH3,3 VH4 | |
1b | ND | 0/8 T cells | —-150 | 0/8 T cells | |||||
ND | 0/10 buffer | —-150 | 0/10 buffer | ||||||
Control | 2′LN | 0/4 T cells | 0/4 T cells | —-150 | 0/4 T cells | ||||
2"LN | 0/6 T cells | —-150 | 0/6 T cells | 0/6 T cells | |||||
2"spleen | 0/5 T cells | —-150 | 0/5 T cells | 0/5 T cells |
Patient No. . | Experiment No. . | Cells Positive for V Rearrangements After PCR With Different Forward Primers . | PCR Products . | Rearrangements . | |||||
---|---|---|---|---|---|---|---|---|---|
Vκ . | VHFRI . | VHFRII . | Clone- Specific . | Total . | Sequenced . | Repeated . | Unique . | ||
1 | 1a | ND | 13/17 | —-150 | ND | 13 | 13 | 13 VH1 | |
1b | ND | 7/14 | —-150 | 7/14 | 7 | 7 | 7 VH1 | ||
2 | 2′LN | 0/13 | 0/13 | —-150 | 3/13 | 3 | 3 | 3 VH1 | |
2"LN | 0/7 | —-150 | 1/7 | 3/7 | 3 | 3 | 3 VH1 | ||
2"spleen | 0/10 | —-150 | 1/10 | 2/10 | 2 | 2 | 2 VH1 | ||
Control | 1a-151 | ND | 6/8 T cells | —-150 | ND | 8 | 8 | 5 VH3,3 VH4 | |
1b | ND | 0/8 T cells | —-150 | 0/8 T cells | |||||
ND | 0/10 buffer | —-150 | 0/10 buffer | ||||||
Control | 2′LN | 0/4 T cells | 0/4 T cells | —-150 | 0/4 T cells | ||||
2"LN | 0/6 T cells | —-150 | 0/6 T cells | 0/6 T cells | |||||
2"spleen | 0/5 T cells | —-150 | 0/5 T cells | 0/5 T cells |
H-RS cells were analyzed together with B and T cells coded in a blinded fashion from two patients with HD. The number of PCR products obtained and sequenced from H-RS and T cells for the first and the second experiment is indicated. Clonally independent VHgene rearrangements of various VH gene families were obtained from control B cells (not shown). Repeated rearrangements indicate clonally related rearrangements; unique rearrangements indicate rearrangements amplified once and not related to any other rearrangement. The sequences obtained from H-RS cells of both patients were deposited in the Genbank under the accession numbers AJ007389 and AJ0007390.
Abbreviations: LN, lymph node; ND, not determined.
PCR was performed either with VHFRI or with VHFRII primers.
In this experiment, a B-cell–specific antibody was eventually used for the “T-cell staining.”