Sosa-Luis Sorely Adelina, Almaraz-Arreortua Alexia, Torres-Aguilar Honorio
Clinical and Basic Immunology Research Department of Biochemical Sciences Faculty, Universidad Autónoma "Benito Juárez" de Oaxaca, Oaxaca, Mexico.
Methods Mol Biol. 2025;2907:299-313. doi: 10.1007/978-1-0716-4430-0_14.
Plasmacytoid dendritic cells (pDCs) are critical in antiviral immune responses and play a significant role in developing autoimmune diseases and transplant rejection. Despite their scarcity in peripheral blood, the accessibility of blood peripheral pDCs makes them a promising source for evaluating T-cell antigen-specific responses in viral infections and autoimmune diseases, as well as alloreactive stimulation in transplantation, including graft-versus-host disease. T-cell priming assays with pDCs have presented a significant challenge due to the low proliferation of T cells. This underscores the critical need for meticulous standardization of suitable conditions for cell purification and coculture in peripheral blood pDC-based mixed leukocyte reactions (MLRs) to ensure productive and quantitative results. This chapter presents a thorough methodology for isolating peripheral blood human pDCs and their application in an MLR assay to evaluate alloreactive T-cell responses using flow cytometry and the adjustments to analyze antigen-specific reactions.
浆细胞样树突状细胞（pDCs）在抗病毒免疫反应中至关重要，并且在自身免疫性疾病的发展和移植排斥反应中发挥重要作用。尽管它们在外周血中数量稀少，但外周血pDCs的可获取性使其成为评估病毒感染和自身免疫性疾病中T细胞抗原特异性反应以及移植中的同种异体反应性刺激（包括移植物抗宿主病）的有前景的来源。由于T细胞增殖率低，用pDCs进行T细胞启动试验面临重大挑战。这突出表明，迫切需要对外周血基于pDC的混合淋巴细胞反应（MLR）中的细胞纯化和共培养的合适条件进行细致的标准化，以确保获得有效且定量的结果。本章介绍了分离外周血人pDCs的详细方法及其在MLR试验中的应用，以使用流式细胞术评估同种异体反应性T细胞反应以及分析抗原特异性反应的调整方法。
