A simple way to increase recovery of the expressed VH and VL genes in single-sorted human B cells
oleh: Sergey V Guselnikov, Tatyana N Belovezhets, Sergey V Kulemzin, Andrey A Gorchakov, Alexander V Taranin
| Format: | Article |
|---|---|
| Diterbitkan: | Taylor & Francis Group 2019-10-01 |
Deskripsi
Cloning VH and VL genes from individual antigen-specific B cells is an attractive approach for producing monoclonal antibodies of the desired specificity. Current RT-PCR protocols, however, result in the successful identification of VH and VL gene pairs in about half of the sorted cells. Here, we demonstrate that single-cell RT-PCR is likely affected by stochastic factors, and that running PCRs in triplicate results in successful amplification of the expressed VH and VL genes in 90–100% of single sorted human B cells.