de StGroth S F, Scheidegger D
J Immunol Methods. 1980;35(1-2):1-21. doi: 10.1016/0022-1759(80)90146-5.
A myeloma line has been developed which produces no globulin chains of its own, has a duplication of 8.7 h, fuses effectively with B-lymphoblasts and produces stable hybrids. An enhancing effect of macrophages on hybridoma yields has been observed. Among the fusing agents tested, PEG of mol.wt. 4000 gave the best results, 20 degrees C being the optimum working temperature. The maintenance medium of choice has been found to be Iscove's with 10% FCS. Direct exposure of fusion cultures to a selective medium with hypoxanthine, aminopterine and thymidine reduced the labor involved and increased the yield. A mechanical device for changing the medium has been designed. The replacement of standard trays by microtrays resulted in a higher frequency of surviving hybrids. By using a feeder layer, the spleen cell input can be reduced 50-fold. At such low multiplicities the positive cultures arise predominantly from single hybrids, eliminating the need for subsequent cloning. The hybrids can be labelled and will yield in serum-free medium. Since at least a third of them inherit the fast growth rate of their myeloma parent and keep producing over 2000 antibody molecules per second, readaptation to ascitic growth is also superfluous. A simplified technique of producing monoclonal antibodies is given in detail, together with the experimental evidence prompting modifications of the classical method of Köhler and Milstein (1975, 1976).
已培育出一种骨髓瘤细胞系，它自身不产生球蛋白链，有8.7小时的复制时间，能与B淋巴细胞有效融合并产生稳定的杂交瘤。已观察到巨噬细胞对杂交瘤产量有增强作用。在所测试的融合剂中，分子量为4000的聚乙二醇效果最佳，20摄氏度是最佳工作温度。已发现最佳的维持培养基是含10%胎牛血清的伊斯科夫培养基。将融合培养物直接置于含次黄嘌呤、氨基蝶呤和胸腺嘧啶的选择培养基中，减少了工作量并提高了产量。已设计出一种更换培养基的机械设备。用微量培养板代替标准培养板可提高杂交瘤存活频率。通过使用饲养层，脾细胞输入量可减少50倍。在如此低的细胞密度下，阳性培养物主要来自单个杂交瘤，无需后续克隆。杂交瘤可进行标记并能在无血清培养基中产生。由于至少三分之一的杂交瘤继承了其骨髓瘤亲本的快速生长速度，每秒能产生2000多个抗体分子，因此也无需再适应腹水生长。详细介绍了一种生产单克隆抗体的简化技术，以及促使对科勒和米尔斯坦经典方法（1975年、1976年）进行改进的实验证据。
