T-cell receptor variable gene analysis of renal allograft-infiltrating cells in biopsy specimens using a nonradioisotopic micromethod
Obata, F.; Kumano, K.; Endo, T.; Takeuchi, Y.; Ohkubo, M.; Tsunoda-Iizuka, M.
Transplantation 66(10): 1389-1392
1998
ISSN/ISBN: 0041-1337 PMID: 9846528 Document Number: 484320
Background. A sensitive micromethod for T-cell receptor (TCR) analysis is needed for clonality analysis of renal allograft-infiltrating T cells (RAITs) obtained by needle biopsy. Methods. TCR cDNA was amplified by the anchored polymerase chain reaction and war. hybridized with 28 different TCR beta variable (TCRBV) genes fixed on nylon membranes, and the percentage of each TCRBV gene was measured spectrophotometrically. Results. The specificity and linearity of the hybridization technique and the constancy of the TCRBV percentages over a wide range of sample amounts were demonstrated by control experiments. Analysis of RAITs of biopsy specimens from four patients showed broad or skewed TCRBV usage, indicating the presence of polyclonal and oligoclonal RAIT populations, respectively. In one patient who received OKT3 immunosuppressive treatment, the TCRBV skewness was dramatically reduced after the treatment. Conclusion. We have established a powerful method for analyzing RAIT clonality, which is especially useful for monitoring RAIT dynamics after immunosuppression therapy.