Kuroki M, Matsumoto Y, Arakawa F, Haruno M, Murakami M, Kuwahara M, Ozaki H, Senba T, Matsuoka Y
First Department of Biochemistry, School of Medicine, Fukuoka University, Japan.
J Immunol Methods. 1995 Mar 13;180(1):81-91. doi: 10.1016/0022-1759(94)00301-c.
To reduce heterophilic antibody interference in a two-site immunoassay for carcinoembryonic antigen (CEA), we utilized a human/mouse chimeric antibody to CEA as the tracer. One mouse monoclonal antibody (MAb), F82-61, which reacts with an epitope present on the domain N of CEA, was immobilized on 96-well polystyrene microtiter plates. A human/mouse chimeric antibody (Ch F11-39), which recognizes an epitope present on the domain B3 of CEA, was biotinylated for the tracer (Ch F11-39 system). Another MAb F11-39, the parental MAb of Ch F11-39, was also biotinylated and used as the control tracer (F11-39 system). For a fair comparison, the same 503 serum samples from healthy individuals were simultaneously assayed in the present study. When a tentative common reference limit of 5 ng/ml was used, the false positive rate with the Ch F11-39 system was only 2.8% (14/503) and that with the F11-39 system was 29.0% (146/503). Adding normal mouse serum (NMS; 1%) or a mixture of purified mouse IgG subclasses (heterophilic blocking reagent (HBR, 15 micrograms/test)) to the F11-39 system reduced the false positive rate from 29.0% to 6.2% (31/503) or 4.8% (24/503), respectively, suggesting that heterophilic antibodies reactive with mouse IgG gave rise to the high positive rate in normal populations with the F11-39 system. On the other hand, the false positive rate with the Ch F11-39 system was only slightly reduced from 2.8% to 2.6% (13/503) or to 2.0% (10/503) by adding NMS or HBR to the Ch F11-39 system. The false positive rates with two commercially available assay systems, CEA Roche EIA.DM or Abbott IMx CEA, were 5.4% (27/503) and 5.8% (29/503), respectively, which both corresponded roughly to that with the F11-39 system including NMS or HBR. These results indicate that the application of human/mouse chimeric antibodies in two-site immunoassays is more effective for reducing interference from heterophilic antibodies than the adding of NMS or purified mouse IgG in the assay using conventional MAbs.
为减少癌胚抗原（CEA）两点免疫分析中嗜异性抗体的干扰，我们使用人/鼠嵌合抗CEA抗体作为示踪剂。一种与CEA N结构域上存在的表位反应的小鼠单克隆抗体（MAb）F82-61被固定在96孔聚苯乙烯微量滴定板上。一种识别CEA B3结构域上存在的表位的人/鼠嵌合抗体（Ch F11-39）被生物素化用作示踪剂（Ch F11-39系统）。另一种MAb F11-39，即Ch F11-39的亲本MAb，也被生物素化并用作对照示踪剂（F11-39系统）。为了进行公平比较，本研究同时检测了来自健康个体的相同503份血清样本。当使用5 ng/ml的暂定共同参考限时，Ch F11-39系统的假阳性率仅为2.8%（14/503），而F11-39系统的假阳性率为29.0%（146/503）。向F11-39系统中加入正常小鼠血清（NMS；1%）或纯化的小鼠IgG亚类混合物（嗜异性阻断试剂（HBR，15微克/检测））分别将假阳性率从29.0%降至6.2%（31/503）或4.8%（24/503），这表明与小鼠IgG反应的嗜异性抗体在F11-39系统的正常人群中导致了高阳性率。另一方面，向Ch F11-39系统中加入NMS或HBR后，Ch F11-39系统的假阳性率仅从2.8%略有降至2.6%（13/503）或2.0%（10/503）。两种市售检测系统，即CEA罗氏EIA.DM或雅培IMx CEA的假阳性率分别为5.4%（27/503）和5.8%（29/503），这两者大致与包括NMS或HBR的F11-39系统的假阳性率相当。这些结果表明，在两点免疫分析中应用人/鼠嵌合抗体比在使用传统MAb的检测中加入NMS或纯化的小鼠IgG更有效地减少嗜异性抗体的干扰。
