Department of Medicine, University of Washington, Seattle, WA, USA.
South African Tuberculosis Vaccine Initiative and Institute of Infectious Disease and Molecular Medicine, Division of Immunology, Department of Pathology, University of Cape Town, Cape Town, South Africa.
J Immunol Methods. 2018 Jul;458:44-52. doi: 10.1016/j.jim.2018.04.004. Epub 2018 Apr 21.
CD1 tetramers loaded with lipid antigens facilitate the identification of rare lipid-antigen specific T cells present in human blood and tissue. Because CD1 proteins are structurally non-polymorphic, these tetramers can be applied to genetically diverse human populations, unlike MHC-I and MHC-II tetramers. However, there are no standardized assays to quantify and characterize lipid antigen-specific T cells present within clinical samples. We incorporated CD1b tetramers loaded with the mycobacterial lipid glucose monomycolate (GMM) into a multi-parameter flow cytometry assay. Using a GMM-specific T-cell line, we demonstrate that the assay is linear, reproducible, repeatable, precise, accurate, and has a limit of detection of approximately 0.007%. Having formally validated this assay, we performed a cross-sectional study of healthy U.S. controls and South African adolescents with and without latent tuberculosis infection (LTBI). We show that GMM-specific T cells are specifically detected in South African subjects with LTBI and not in U.S. healthy controls. This assay can be expanded to include additional tetramers or phenotypic markers to characterize GMM-specific T cells in studies of mycobacterial infection, disease, or vaccination.
CD1 四聚体负载脂质抗原，有助于鉴定存在于人体血液和组织中的稀有脂质抗原特异性 T 细胞。由于 CD1 蛋白在结构上是非多态性的，与 MHC-I 和 MHC-II 四聚体不同，这些四聚体可以应用于遗传多样性的人群。然而，目前还没有标准化的检测方法来定量和描述临床样本中存在的脂质抗原特异性 T 细胞。我们将负载分枝杆菌脂质葡萄糖单酰基（GMM）的 CD1b 四聚体纳入多参数流式细胞术检测中。使用 GMM 特异性 T 细胞系，我们证明该检测方法是线性的、可重复的、可重现的、精确的、准确的，检测限约为 0.007%。该检测方法经过正式验证后，我们对美国健康对照组和南非潜伏性结核感染（LTBI）青少年进行了横断面研究。我们发现，LTBI 的南非受试者中可特异性检测到 GMM 特异性 T 细胞，而美国健康对照组中则没有。该检测方法可以扩展到包括其他四聚体或表型标志物，以在分枝杆菌感染、疾病或疫苗接种的研究中对 GMM 特异性 T 细胞进行特征描述。
