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Abstract：
It is generally recognized that dysregulation of the immune system plays a critical role in many diseases, including autoimmune diseases and cancer. T cells play a crucial role in maintaining self-tolerance, while loss of immune tolerance and T cell activation can lead to severe inflammation and tissue damage. T cell responses have a key role in the effectiveness of vaccination strategies and immunomodulating therapies. Immunomonitoring methods have the ability to elucidate immunological processes, monitor the development of disease and assess therapeutic effects. In this respect, it is of particular interest to evaluate antigen (Ag)-specific T cells by determining their frequency, type and functionality in cellular assays. Nevertheless, Ag-specific T cells are detected infrequently in most diseases using current techniques. Many efforts have been made to develop more sensitive, reproducible, and reliable methods for Ag-specific T cell detection. It has been found that analysis of cellular proliferation can be a useful tool to determine the presence and frequency of Ag-specific T cell and to provides insight into modulation of the T cell response by a specific antigen or therapy. However, the selection of a cut-off value for a positive response and therefore a more accurate interpretation of the data, continues to be a major concern. Here, we provide guidelines to select a proper cut-off for monitoring of Ag-specific CD4+ T cell responses. In vitro Ag-stimulation has been assessed with two methods; a dye-based proliferation assay and 3H-thymidine-based assay. Two cut-off approaches are compared; mean and variance of control wells, and the stimulation index. By evaluating the proliferative response to the in vitro Ag-stimulation using these two methods, we demonstrate the importance of taking into consideration the variability of the control wells to distinguish a positive from a false positive response.
摘要：
人们普遍认为，免疫系统的失调在许多疾病中起着至关重要的作用。包括自身免疫性疾病和癌症。T细胞在维持自我耐受中起着至关重要的作用。而免疫耐受和T细胞活化的丧失可导致严重的炎症和组织损伤。T细胞应答在疫苗接种策略和免疫调节疗法的有效性中具有关键作用。免疫监测方法具有阐明免疫过程的能力，监测疾病的发展并评估治疗效果。在这方面,特别感兴趣的是通过确定它们的频率来评估抗原（Ag）特异性T细胞，细胞测定中的类型和功能。然而，使用当前技术在大多数疾病中很少检测到Ag特异性T细胞。已经做出了许多努力来发展更敏感的，可重复,和可靠的Ag特异性T细胞检测方法。已经发现,细胞增殖的分析可以是确定Ag特异性T细胞的存在和频率并提供对特异性抗原或疗法对T细胞应答的调节的洞察的有用工具。然而,选择阳性响应的截止值，从而对数据进行更准确的解释，仍然是一个主要问题。这里,我们提供指导,以选择适当的截止值来监测Ag特异性CD4+T细胞应答.已经通过两种方法评估了体外Ag刺激;基于染料的增殖测定和基于3H-胸苷的测定。比较了两种截止方法；对照井的均值和方差，和刺激指数。通过使用这两种方法评估对体外Ag刺激的增殖反应，我们证明了考虑对照孔的变异性以区分阳性和假阳性的重要性。
