Inouye H, Hank J A, Chardonnens X, Segall M, Alter B J, Bach F H
J Exp Med. 1980 Aug 1;152(2 Pt 2):143s-155s.
Human T lymphocytes obtained as blasts on day 4 from a primary mixed leukocyte culture (MLC) were cloned in the presence of T cell growth factor (TCGF) and feeder cells. Parameters important in producing higher-specific-activity TCGF were evaluated; irradiation of the responding cells as well as removal of adherent cells or inclusion of indomethacin in the culture was important. In addition, the presence of an irradiated lymphoblastoid cell line (LCL) cell in the TCGF-producing system enhanced activity in the supernate. The long-term maintenance of progeny from clones was achieved by utilizing the LCL autologous with either the responding or sensitizing cells from the initial MLC as feeder cells. Under those conditions, clones could be expanded for 7 or more wk with the maintenance of PLT reactivity. Had all the cells in each clone been maintained for the full 7 wk, more than 1 X 10(10) cells could have been developed in each clone. The cloned reagents provide a higher degree of antigen-specific reactivity than do normal PLT cells. It is to be anticipated that as the requirements for cloning are made more stringent, including the recloning of the cells, these reagents will aid greatly in the dissection of the complexity attendant to HLA-D.
从初次混合淋巴细胞培养（MLC）第4天获得的作为母细胞的人T淋巴细胞，在T细胞生长因子（TCGF）和饲养细胞存在的情况下进行克隆。对产生更高比活性TCGF重要的参数进行了评估；照射反应细胞以及去除贴壁细胞或在培养物中加入消炎痛很重要。此外，在产生TCGF的系统中存在经照射的淋巴母细胞系（LCL）细胞可增强上清液中的活性。通过利用与初始MLC中的反应细胞或致敏细胞自体的LCL作为饲养细胞，实现了克隆后代的长期维持。在这些条件下，克隆可扩增7周或更长时间，并维持PLT反应性。如果每个克隆中的所有细胞都维持完整的7周，每个克隆中可能会产生超过1×10¹⁰个细胞。与正常PLT细胞相比，克隆试剂提供了更高程度的抗原特异性反应性。可以预期，随着克隆要求变得更加严格，包括细胞的再克隆，这些试剂将极大地有助于剖析与HLA-D相关的复杂性。
