Bankert R B, Mazzaferro D, Mayers G L
Hybridoma. 1981;1(1):47-58. doi: 10.1089/hyb.1.1981.1.47.
Hybridoma clones producing anti-hapten antibodies were established by fusing spleen cells from mice immunized with 4-azophthalate keyhole limpet hemocyanin to drug-resistant non-producing myelomas. The affinity and homogeneity of the immunochemically purified anti-phthalate antibodies were determined by equilibrium dialysis. We report here a broad distribution of the monoclonal antibody affinities ranging from a low of 4.0 X 10(4) to a high of 4.0 X 10(7) (L/M). The binding data for eleven anti-phthalate antibodies described a straight line in a Scatchard plot as would be expected for homogeneous antibodies. We determined that all hybridomas, except those secreting very low affinity antibody, produced hemolytic plaques. The hybridomas made it possible to test several assumptions regarding the association of plaque morphology and plaque inhibition with the affinity of the antibody secreted by the plaque-forming cells. Our studies indicate that the affinity of the antibody secreted by the hybridoma clones does not correlate with either the size of the plaque or with the efficiency with which the hybridoma-produced plaques are inhibited by free hapten. By comparing hybridoma-produced plaques to plaques produced by phthalate-immune spleen cells it has been demonstrated that the plaque size within each hybridoma clone was substantially less heterogeneous than that observed for the immune spleen cells. Our results do support the assumption that the range of free hapten concentration over which PFC are inhibited is a reflection of PFC heterogeneity. The analysis of the PFC inhibition of hybridomas produced an interesting and unexpected result which is reported here. It was observed that subinhibitory doses of free hapten caused an enhancement in the number of hybridoma-produced plaques. The relevance of this finding to the recent observation and interpretation of plaque enhancement (i.e., the displacement of anti-idiotype antibodies) observed for immune spleen cells is discussed.
通过将用4-偶氮邻苯二甲酸钥孔戚血蓝蛋白免疫的小鼠脾细胞与耐药性非分泌型骨髓瘤细胞融合，建立了产生抗半抗原抗体的杂交瘤克隆。通过平衡透析法测定了免疫化学纯化的抗邻苯二甲酸抗体的亲和力和均一性。我们在此报告单克隆抗体亲和力的广泛分布，范围从低至4.0×10⁴到高至4.0×10⁷（L/M）。11种抗邻苯二甲酸抗体的结合数据在Scatchard图中呈直线，这是均一抗体所预期的。我们确定，除了那些分泌极低亲和力抗体的杂交瘤外，所有杂交瘤都产生溶血空斑。这些杂交瘤使得能够测试关于空斑形态和空斑抑制与形成空斑细胞分泌的抗体亲和力之间关联的几个假设。我们的研究表明，杂交瘤克隆分泌的抗体亲和力与空斑大小或游离半抗原抑制杂交瘤产生的空斑的效率均无相关性。通过将杂交瘤产生的空斑与邻苯二甲酸免疫脾细胞产生的空斑进行比较，已证明每个杂交瘤克隆内的空斑大小比免疫脾细胞所观察到的要少得多的异质性。我们的结果确实支持这样的假设，即抑制PFC的游离半抗原浓度范围反映了PFC的异质性。对杂交瘤的PFC抑制分析产生了一个有趣且出乎意料的结果，在此报告。观察到亚抑制剂量的游离半抗原导致杂交瘤产生的空斑数量增加。讨论了这一发现与最近观察到的免疫脾细胞空斑增强（即抗独特型抗体的置换）及其解释的相关性。
