Novel method to detect T clonality and the antigen specificity

为了鉴定抗原特异性T细胞应答的存在并跟踪这些反应的变化，评估某些T细胞是否在淋巴细胞群中克隆积累被认为是有用的。为此，我们已经建立了一种新的方法来分析T细胞克隆性使用相结合的逆转录酶-聚合酶链反应的T细胞受体β链转录本和单链构象多态性（SSCP）。使用这种方法，我们从异质性T细胞群体中获得了涂片样的DNA。另一方面，单个T细胞克隆在适当的V β扩增中显示一条带，异质淋巴细胞群中的累积T细胞克隆被鉴定为背景涂片模式中的一条带。如果淋巴细胞群体在体外或体内被抗原刺激，则在背景涂片中发现产生几条不同的条带。因此，该方法可用于监测T细胞克隆反应的动态变化。对包括自身免疫性疾病、恶性疾病和移植在内的几种免疫性疾病的分析揭示了抗原特异性T细胞免疫应答在这些疾病中的参与。此外，利用SSCP凝胶条带的重复性，我们现在能够比较不同样品中积累的T细胞克隆的身份，而不需要对每个克隆进行核苷酸测序。因此，这些信息可以阐明宿主中均匀或稳定的免疫反应的发生，并且还表明这些反应在体内起重要作用。因此，总的来说，上述新型T细胞克隆性分析被认为可用于研究免疫学各个领域中的T细胞免疫应答。

To identity the existence of antigen-specific T cell responses and to follow the changes of these reactions, it is considered useful to evaluate whether certain T cells clonally accumulate in the lymphocyte population. For this purpose, we have established a novel method to analyze T cell clonality using a combination of reverse transcriptase-polymerase chain reaction of T cell receptor beta chain transcripts and single-strand conformation polymorphism (SSCP). Using this method, we obtained a smear-like pattern of electrophoresed DNA from the heterogeneous T cell population. On the other hand, a single T cell clone exhibits a band in appropriate Vbeta amplification and an accumulatedT cell clone in a heterogeneous lymphocyte population is identified as a band in the background smear pattern. If a lymphocyte population was stimulated by an antigen either in vitro or in vivo, several distinct bands were found to be generated in the background smear. Thus, the dynamic changes of T cell clonal responses could be monitored with this method. Analyzes of several immunological disorders, including autoimmune diseases, malignant disorders, andtransplantation, revealed the involvement of antigen-spesific T cell immune responses in these disorders. Furthermore, taking advantege of the reproduciblemobility of a band of SSCP gel, we are now able tocompare identities of the accumulated T cell Clones in different samples without the need for nucleoptidesequencing of each clone. Such information can thus elucidate the occurrence of uniform or stable immunological reactions in the host and also suggests that these reactions play an important role in vivo. Therefore, taken togeter, the above described novel T cell clonality analysis is considered to be useful in studying the T cell immune responses in various fields of immunology.