Emory Vaccine Center at the Yerkes National Primate Center at Emory University, United States.
J Immunol Methods. 2012 Jan 31;375(1-2):118-28. doi: 10.1016/j.jim.2011.09.016. Epub 2011 Oct 8.
Detection of antigen-specific T cells at the single-cell level by ELISpot or flow cytometry techniques employing intracellular cytokine staining (ICS) is now an indispensable tool in many areas of immunology. When precisely mapped, optimal MHC-binding peptide epitopes are unknown, these assays use antigen in a variety of forms, including recombinant proteins, overlapping peptide sets representing one or more target protein sequences, microbial lysates, lysates of microbially-infected cells, or gene delivery vectors such as DNA expression plasmids or recombinant vaccinia or adenoviruses expressing a target protein of interest. Here we introduce replication-restricted, recombinant vesicular stomatitis virus (VSV) vectors as a safe, easy to produce, simple to use, and highly effective vector for genetic antigen delivery for the detection of human antigen-specific helper and cytotoxic T cells. To demonstrate the broad applicability of this approach, we have used these vectors to detect human T cell responses to the immunodominant pp65 antigen of human cytomegalovirus, individual segments of the yellow fever virus polyprotein, and to various influenza proteins.
通过酶联免疫斑点法（ELISpot）或流式细胞术技术，利用细胞内细胞因子染色（ICS）在单细胞水平上检测抗原特异性 T 细胞，现在已成为免疫学许多领域不可或缺的工具。当精确绘制时，最佳 MHC 结合肽表位是未知的，这些检测使用各种形式的抗原，包括重组蛋白、代表一个或多个靶蛋白序列的重叠肽集、微生物裂解物、微生物感染细胞的裂解物或基因传递载体，如 DNA 表达质粒或表达感兴趣的靶蛋白的重组单纯疱疹病毒或腺病毒。在这里，我们介绍复制受限的重组水疱性口炎病毒（VSV）载体，作为一种安全、易于生产、易于使用且高效的基因抗原传递载体，用于检测人类抗原特异性辅助和细胞毒性 T 细胞。为了证明这种方法的广泛适用性，我们已经使用这些载体来检测人类 T 细胞对人巨细胞病毒的免疫优势 pp65 抗原、黄热病毒多蛋白的各个片段以及各种流感蛋白的反应。
