Malaise M G, Franchimont P, Mahieu P R
Department of Internal Medicine, State University of Liège, Belgium.
J Immunol Methods. 1989 May 12;119(2):231-9. doi: 10.1016/0022-1759(89)90401-8.
The percentage of normal human monocytes (MCs) that are able to form rosettes with, and subsequently phagocytose, IgG-coated red blood cells (RBCs) has been determined in vitro using five batches of anti-RBC IgG antibodies. These antibodies differed from each other by their capacity to bind to lectins recognizing two of the oligosaccharide structures of the Fc domain, namely, peanut agglutinin (PNA) and concanavalin A (ConA) which specifically bind to beta-galactosyl and alpha-mannosyl residues, respectively. The threshold between high (H) and low (L) binding capacities (BC) was arbitrarily fixed at 15% of mean specific binding. For each level of RBC sensitization tested (1500-6000 Ab molecules/one RBC), the percentage of MCs binding at least three IgG-RBCs was similar whatever the IgG Ab preparations used. In contrast, the percentage of MCs capable of phagocytosing at least three IgG-RBCs coated with 3000, 4500 and 6000 IgG/cell, as well as the phagocytosis index (number of IgG-RBCs ingested/100 MCs) of IgG-RBCs coated with 1500, 3000, 4500 and 6000 IgG/cell, were significantly lower (P less than 0.01 at least) using IgG Ab molecules with either [(PNA-H)(ConA-H)] BC, [(PNA-L) (ConA-H)] BC or with [(PNA-L)(ConA-L)] BC than the corresponding values measured using RBCs coated with IgG Ab molecules exhibiting [(PNA-H)(ConA-L)] BC. The binding to MCs of 125I-labelled anti-RBC IgG Ab molecules exhibiting different binding profiles to PNA and to ConA was studied by Scatchard plot analysis. A single class of binding sites was observed in each case. MCs bound a mean of 23,000 IgG molecules with a mean association constant (Ka) for IgG binding of about 1.4 X 10(8) M-1. These data indicate that terminal (and/or accessible) galactosyl and mannosyl residues of IgG Ab molecules play a role in the ingestion of IgG-RBCs by human MCs, despite the fact IgG Ab binding to IgG(Fc) receptors is not significantly affected. Thus, when studying the phagocytosis of IgG-coated RBC by human MC monolayers, the assay should be performed not only using similar RBC/MC ratios and IgG coating values, but also with IgG antibodies having comparable mean PNA and ConA binding capacities.
使用五批抗红细胞IgG抗体，在体外测定了能够与IgG包被的红细胞（RBC）形成玫瑰花结并随后吞噬它们的正常人单核细胞（MC）的百分比。这些抗体彼此的区别在于它们与识别Fc结构域两种寡糖结构的凝集素结合的能力，即花生凝集素（PNA）和伴刀豆球蛋白A（ConA），它们分别特异性结合β-半乳糖基和α-甘露糖基残基。高（H）结合能力（BC）和低（L）结合能力之间的阈值被任意设定为平均特异性结合的15%。对于测试的每个红细胞致敏水平（1500 - 6000个抗体分子/一个红细胞），无论使用何种IgG抗体制剂，结合至少三个IgG - RBC的单核细胞百分比相似。相比之下，使用具有[(PNA - H)(ConA - H)] BC、[(PNA - L)(ConA - H)] BC或[(PNA - L)(ConA - L)] BC的IgG抗体分子时，能够吞噬至少三个每细胞包被3000、4500和6000个IgG的IgG - RBC的单核细胞百分比，以及每细胞包被1500、3000、4500和6000个IgG的IgG - RBC的吞噬指数（摄入的IgG - RBC数量/100个单核细胞），均显著低于（至少P小于0.01）使用具有[(PNA - H)(ConA - L)] BC的IgG抗体分子包被的红细胞所测得的相应值。通过Scatchard图分析研究了对PNA和ConA呈现不同结合谱的125I标记的抗红细胞IgG抗体分子与单核细胞的结合。在每种情况下均观察到单一类别的结合位点。单核细胞平均结合23,000个IgG分子，IgG结合的平均缔合常数（Ka）约为1.4×108 M-1。这些数据表明，IgG抗体分子的末端（和/或可及的）半乳糖基和甘露糖基残基在人单核细胞摄取IgG - RBC中起作用，尽管IgG抗体与IgG（Fc）受体的结合未受到显著影响。因此，在研究人单核细胞单层对IgG包被的红细胞的吞噬作用时，不仅应使用相似的红细胞/单核细胞比例和IgG包被值进行测定，还应使用具有可比的平均PNA和ConA结合能力的IgG抗体。
