Utsunomiya Y, Bill J, Palmer E, Gollob K, Takagaki Y, Kanagawa O
Lilly Research Laboratories, La Jolla, CA 92037.
J Immunol. 1989 Oct 15;143(8):2602-8.
Three rat mAb, RR3-15, RR3-16, and RR3-18, were established by fusing spleen cells from a rat immunized with the male Ag-specific cytolytic T cell clone, OH6, to mouse myeloma cells. The mAb was identified by their capacity to focus the cytolytic activity of the OH6 CTL clone on nonspecific target cells via FcR-FcR interaction. That all three mAb recognized the OH6 TCR was confirmed by immunoprecipitation studies in which each antibody precipitated a 90 kDa disulfide-linked heterodimer characteristic of the TCR. Surface immunofluorescence staining of a panel of T cell lines and splenic T cell populations showed that RR3-16 reacted not only to the OH6 T cell clone but also to a minor fraction of normal T cells. This reactivity was found to be due to the expression of a gene in the V alpha 3 family. However, RR3-16 did not react with all T cell lines and clones known to express genes from the V alpha 3 family. cDNA sequences of three independent RR3-16+ T cell hybridomas analyzed by polymerase chain reaction were identical to the previously published V alpha 3 sequence of the CTL clone C9. Thus, the mAb RR3-16 is specific for a single member of the TCR V alpha 3 gene family. Analysis of the expression of RR3-16+ TCR in CD4+ and CD8+ subsets of peripheral T cells demonstrated preferential expression on CD8+ T cells, suggesting regulated expression of this particular TCR V alpha gene.
通过将用雄性抗原特异性细胞溶解T细胞克隆OH6免疫的大鼠的脾细胞与小鼠骨髓瘤细胞融合，建立了三种大鼠单克隆抗体RR3-15、RR3-16和RR3-18。通过其将OH6 CTL克隆的细胞溶解活性通过FcR-FcR相互作用聚焦于非特异性靶细胞的能力来鉴定该单克隆抗体。免疫沉淀研究证实所有三种单克隆抗体都识别OH6 TCR，其中每种抗体都沉淀出一种90 kDa的二硫键连接的异二聚体，这是TCR的特征。一组T细胞系和脾T细胞群体的表面免疫荧光染色显示，RR3-16不仅与OH6 T细胞克隆反应，还与一小部分正常T细胞反应。发现这种反应性是由于Vα3家族中一个基因的表达。然而，RR3-16并不与所有已知表达Vα3家族基因的T细胞系和克隆反应。通过聚合酶链反应分析的三个独立的RR3-16 + T细胞杂交瘤的cDNA序列与先前发表的CTL克隆C9的Vα3序列相同。因此，单克隆抗体RR3-16对TCR Vα3基因家族的单个成员具有特异性。对外周T细胞的CD4 +和CD8 +亚群中RR3-16 + TCR表达的分析表明，其在CD8 + T细胞上优先表达，提示该特定TCR Vα基因的表达受到调控。
