Measurement and Clinical Monitoring of Human Lymphocyte Clonality by Massively Parallel V-D-J Pyrosequencing

作者：Scott Dexter Boyd, Eleanor L. Marshall, Jason D. Merker, Jay M. Maniar, Lyndon N. Zhang, Bita Sahaf, Carol D. Jones, Birgitte B. Simen, Bozena Hanczaruk, Khoa D. Nguyen, Kari Christine Nadeau, Michael Egholm, David Bernard Miklos, James L. Zehnder, Andrew Z. Fire · 发表于：Science Translational Medicine · 年份：2009 · DOI：10.1126/scitranslmed.3000540 · 被引用次数：400 · 研究领域：T-cell and B-cell Immunology、Immunotherapy and Immune Responses、Single-cell and spatial transcriptomics

The complex repertoire of immune receptors generated by B and T cells enables recognition of diverse threats to the host organism. In this work, we show that massively parallel DNA sequencing of rearranged immune receptor loci can provide direct detection and tracking of immune diversity and expanded clonal lymphocyte populations in physiological and pathological contexts. DNA was isolated from blood and tissue samples, a series of redundant primers was used to amplify diverse DNA rearrangements, and the resulting mixtures of barcoded amplicons were sequenced using long-read ultra deep sequencing. Individual DNA molecules were then characterized on the basis of DNA segments that had been joined to make a functional (or nonfunctional) immune effector. Current experimental designs can accommodate up to 150 samples in a single sequence run, with the depth of sequencing sufficient to identify stable and dynamic aspects of the immune repertoire in both normal and diseased circumstances. These data provide a high-resolution picture of immune spectra in normal individuals and in patients with hematological malignancies, illuminating, in the latter case, both the initial behavior of clonal tumor populations and the later suppression or re-emergence of such populations after treatment.