Lewis G K, Goodman J W
J Exp Med. 1978 Oct 1;148(4):915-24. doi: 10.1084/jem.148.4.915.
Strain A/J mice immunized with azobenzenearsonate (ABA)-mouse IgG conjugates develop suppression for anti-trinitrophenyl(TNP) responses to doubly conjugated (ABA,TNP) proteins. This suppression is specific for the ABA epitope and is mediated by T cells in cell transfer experiments. ABA-binding T cells from suppressed animals were purified by a two-stage procedure in which B cells were removed from spleen cell populations by adherence to plastic surfaces coated with anti-mouse Ig antibody, followed by binding the nonadherent population (more 95 percent Thy-1-positive) to surfaces coated with ABA-protein conjugates. Approximately 90 percent of the cells recovered by temperature-dependent elution from the ABA plates (similar to 2 percent of the spleen cells) bound antigen immediately afterward, and up to 50 percent of the cells bound anti-cross-reactive idiotype antibody. On the other hand, the nonadherent T-cell population was completely negative in the antigen- binding and idiotype assays. Another distinguishing feature of the two T-cell populations was that 78 percent of the adherent cells, but only 2 percent of the nonadherent cells, were Ia positive, although the specific I-region marker(s) expressed on the cells was not identified. The biological function of the antigen-binding T cells was investigated using a standard cell transfer protocol. Suppressor cells were enriched in the adherent population by a factor of at least 25, establishing that functional, epitope-specific, idiotype-bearing T cells can be significantly purified by this procedure. Note Added in Proof. We have recently isolated two types of ABA-binding molecules biosynthetically labeled with (35)S-methionine from NP-40 lysates of purified antigen-specific T cells. The molecules were purified by adsorption onto an ABA-Sepharose immunoadsorbent followed by elution with 9 M urea. Autoradiograms of SDS-PAGE of the eluates revealed components with tool wt of approximately 60,000 and 33,000 dahons. These molecules were not present in eluates from a bovine IgG-Sepharose control immunoadsorbent and thus represent specific ABA-binding products synthesized by T cells.
用偶氮苯砷酸盐（ABA）-小鼠IgG缀合物免疫的A/J品系小鼠，对双缀合（ABA，三硝基苯基[TNP]）蛋白的抗三硝基苯基（TNP）反应产生抑制作用。这种抑制作用对ABA表位具有特异性，并且在细胞转移实验中由T细胞介导。通过两步法纯化来自受抑制动物的ABA结合T细胞，其中通过粘附于包被有抗小鼠Ig抗体的塑料表面从脾细胞群体中去除B细胞，然后将非粘附群体（超过95%为Thy-1阳性）与包被有ABA-蛋白缀合物的表面结合。通过温度依赖性洗脱从ABA平板上回收的细胞（约占脾细胞的2%）中，约90%的细胞随后立即结合抗原，高达50%的细胞结合抗交叉反应独特型抗体。另一方面，非粘附T细胞群体在抗原结合和独特型检测中完全呈阴性。这两种T细胞群体的另一个显著特征是，78%的粘附细胞呈Ia阳性，但只有2%的非粘附细胞呈Ia阳性，尽管未鉴定出细胞上表达的特定I区标记。使用标准的细胞转移方案研究了抗原结合T细胞的生物学功能。抑制细胞在粘附群体中的富集倍数至少为25倍，证实通过该方法可以显著纯化功能性、表位特异性、携带独特型的T细胞。 补充证明。我们最近从纯化的抗原特异性T细胞的NP-40裂解物中分离出两种用（35）S-甲硫氨酸进行生物合成标记的ABA结合分子。这些分子通过吸附到ABA-琼脂糖免疫吸附剂上，然后用9M尿素洗脱进行纯化。洗脱物的SDS-PAGE放射自显影片显示分子量约为60,000和33,000道尔顿的组分。这些分子不存在于牛IgG-琼脂糖对照免疫吸附剂的洗脱物中，因此代表T细胞合成的特异性ABA结合产物。
