Pierce C W, Johnson B M, Gershon H E, Asofsky R
J Exp Med. 1971 Aug 1;134(2):395-416. doi: 10.1084/jem.134.2.395.
We have demonstrated for the first time that mouse spleen cells stimulated in vitro with heterologous erythrocytes developed immunoglobulin class-specific gammaM, gamma(1), gamma(2a+2b), and gammaA plaque-forming cell (PFC) responses. A modification of the hemolytic plaque technique, the addition of goat anti-mouse micro-chain antibody to the assay preparation, specifically prevented development of all gammaM PFC and enabled accurate and reproducible enumeration of immunoglobulin class-specific PFC after treatment with appropriate monospecific anti-globulins and complement. Culture conditions, with regard to medium, atmosphere, agitation, and spleen cell densities, were similar to those previously shown to support only gammaM PFC responses. Evaluation of the kinetics of appearance of PFC showed that gammaM PFC reached maximum numbers on days 4-5; the magnitude of this response was 3-10 times greater than gamma(1) gamma(2a+2b), or gammaA PFC which reached maximum numbers on days 5-6. Optimal erythrocyte antigen dose for gammaM PFC responses was 10(7)/culture, whereas a dose of 10(6) erythrocytes/culture consistently stimulated optimal gamma(1) gamma(2a+2b), or gammaA PFC responses. Investigations of the effects of anti-erythrocyte antibody on gammaM and gammaG PFC responses indicated that antibody suppressed these responses by neutralizing the effective antigenic stimulus at the macrophage-dependent phase of the response. At the same antibody concentration, gammaG PFC responses were more effectively suppressed than gammaM PFC responses. Further, gammaG responses could be almost completely suppressed by antibody as long as 48 hr after initiation of cultures, whereas gammaM PFC responses could only be completely suppressed during the first 24 hr. These results were discusssed in terms of the role of antigen in the stimulation gammaM and gammaG antibody.
我们首次证明,用异源红细胞在体外刺激的小鼠脾细胞产生了免疫球蛋白类特异性γM、γ(1)、γ(2a + 2b)和γA空斑形成细胞(PFC)反应。溶血空斑技术的一种改进方法,即在检测制剂中添加山羊抗小鼠微链抗体,特异性地阻止了所有γM PFC的产生,并在用适当的单特异性抗球蛋白和补体处理后,能够准确且可重复地计数免疫球蛋白类特异性PFC。关于培养基、气体环境、搅拌和脾细胞密度的培养条件,与先前显示仅支持γM PFC反应的条件相似。对PFC出现动力学的评估表明,γM PFC在第4 - 5天达到最大数量;该反应的强度比γ(1)、γ(2a + 2b)或γA PFC大3 - 10倍,γ(1)、γ(2a + 2b)或γA PFC在第5 - 6天达到最大数量。γM PFC反应的最佳红细胞抗原剂量为10(7)/培养物,而10(6)个红细胞/培养物的剂量始终能刺激最佳的γ(1)、γ(2a + 2b)或γA PFC反应。对抗红细胞抗体对γM和γG PFC反应影响的研究表明,抗体通过在反应的巨噬细胞依赖阶段中和有效的抗原刺激来抑制这些反应。在相同抗体浓度下,γG PFC反应比γM PFC反应更有效地受到抑制。此外,只要在培养开始后48小时内,γG反应几乎可以被抗体完全抑制,而γM PFC反应只能在最初24小时内被完全抑制。根据抗原在刺激γM和γG抗体中的作用对这些结果进行了讨论。