Quantitative analysis of T cell receptor diversity in clinical samples of human peripheral blood

The analysis of T cell receptor diversity provides a clinically relevant and sensitive marker of repertoire loss, gain, or skewing. Spectratyping is a broadly utilized technique to measure global TCR diversity by the analysis of the lengths of CDR3 fragments in each V beta family. However the common use of large numbers of T cells to obtain a global view of TCR V beta CDR3 diversity has restricted spectratyping analyses when limited T-cell numbers are available in clinical setting, such as following transplant regimens. We here demonstrate that one hundred thousand T cells are sufficient to obtain a robust, highly reproducible measure of the global TCR V beta repertoire diversity among twenty V beta families in human peripheral blood. We also show that use of lower cell number results not in a dwindling of observed diversity but rather in non-reproducible patterns in replicate spectratypes. Finally, we report here a simple to use but sensitive method to quantify repertoire divergence in patient samples by comparison to a standard repertoire profile we generated from fifteen normal donors. We provide examples using this method to statistically evaluate the changes in the global TCR V beta repertoire diversity that may take place during T subset immune reconstitution after hematopoietic stem cell transplantation or after immune modulating therapies. Published by Elsevier B.V.