Czop J K, Gurish M F, Kadish J L
Department of Medicine, Harvard Medical School, Boston, MA.
J Immunol. 1990 Aug 1;145(3):995-1001.
beta-Glucan receptors are present on mammalian phagocytic cells and provide an important physiologic mechanism for opsonin-independent clearance of yeasts and fungi. To prepare an immunologic probe to human monocyte beta-glucan receptors, an approach was taken that focused on the ligand specificity of the receptors as expressed by an anti-Id. The algal beta-glucan laminarin was chemically coupled to protein carriers to prepare an immunogenic beta-glucan. Three mouse IgG2a mAb were raised against laminarin, and one, mAb OEA10, exhibited specificity for the soluble unit ligand yeast heptaglucoside and the ligands present on zymosan and glucan particles that are recognized by monocyte beta-glucan receptors. These findings prompted usage of mAb OEA10 as immunogen for the preparation of an anti-Id. The resulting rabbit antiserum was subjected to sequential immunoaffinity chromatography to purify anti-idiotypic antibodies. The final product contained only IgG by SDS-PAGE and was shown to be specific by its selectively blocking binding of 125I-mAb OEA10 to laminarin. Pretreatment of adherent monocytes with 0.4 micrograms/ml of the anti-Id reduced the numbers of monocytes ingesting zymosan and glucan particles by 64 and 43%, respectively, whereas ingestion of IgG-coated SRBC was unaffected by as much as 16 micrograms/ml of the anti-Id. Incubation of adherent monocytes with increasing amounts of 125I-anti-Id in the absence and presence of 40-fold molar excess unlabeled anti-Id revealed dose-dependent specific binding, which approached plateau levels with 1 microgram/ml of labeled anti-Id. Thus, the anti-Id binds to monocytes and displays functional characteristics of soluble beta-glucan ligands, indicating that some of the anti-idiotypic antibodies recognize epitopes within monocyte beta-glucan receptors.
β-葡聚糖受体存在于哺乳动物吞噬细胞上，为酵母和真菌的非调理素依赖性清除提供了重要的生理机制。为了制备针对人单核细胞β-葡聚糖受体的免疫探针，采用了一种关注抗独特型抗体所表达的受体配体特异性的方法。将藻类β-葡聚糖海带多糖化学偶联到蛋白质载体上，制备免疫原性β-葡聚糖。针对海带多糖产生了三种小鼠IgG2a单克隆抗体，其中一种单克隆抗体OEA10对可溶性单位配体酵母七糖以及被单核细胞β-葡聚糖受体识别的酵母聚糖和葡聚糖颗粒上的配体具有特异性。这些发现促使使用单克隆抗体OEA10作为免疫原制备抗独特型抗体。对所得兔抗血清进行连续免疫亲和层析以纯化抗独特型抗体。通过SDS-PAGE分析，最终产物仅含有IgG，并且通过其选择性阻断125I-单克隆抗体OEA10与海带多糖的结合而显示出特异性。用0.4微克/毫升的抗独特型抗体预处理贴壁单核细胞，可使摄取酵母聚糖和葡聚糖颗粒的单核细胞数量分别减少64%和43%，而摄取IgG包被的绵羊红细胞则不受高达16微克/毫升抗独特型抗体的影响。在不存在和存在40倍摩尔过量未标记抗独特型抗体的情况下，用越来越多的125I-抗独特型抗体孵育贴壁单核细胞，显示出剂量依赖性特异性结合，当标记抗独特型抗体浓度为1微克/毫升时接近平台水平。因此，抗独特型抗体与单核细胞结合并表现出可溶性β-葡聚糖配体的功能特性，表明一些抗独特型抗体识别单核细胞β-葡聚糖受体内的表位。
