Deftos L J, Wolfert R L, Hill C S, Burton D W
Department of Medicine, San Diego VA Medical Center, University of California.
Clin Chem. 1992 Nov;38(11):2318-21.
We developed a panel of monoclonal antibodies to human bone gla protein (BGP; osteocalcin) peptides that span the linear sequence of the molecule, specifically BGP 1-12 (N-terminal), BGP 15-30 (midregion), and BGP 38-49 (C-terminal). These antibodies were evaluated in various combinations of two-site formats in studies of serum BGP concentrations. For clinical studies, we selected from a panel of antibodies the two most sensitive antibody pairs for the intact molecule (N-C); we also used a polyclonal RIA based on BGP-C. For the two-site format, we used two N-terminal antibodies, 029 and 052, adsorbed to polystyrene beads, and radioiodinated a C-terminal antibody, 663. The standard for each of the assays was purified human BGP. The following BGP serum concentrations (microgram/L, mean +/- SE) were measured with the various assays: by the 029-663 assay, results for normal subjects were 7 +/- 3, for patients with renal failure 25 +/- 8, and for patients with Paget disease 12 +/-4; by the 052-663 assay, the respective results were 22 +/- 4, 44 +/- 12, and 31 +/- 7; by the polyclonal assay, the results were 3 +/- 0.2, 13 +/- 2, and 5 +/- 1. The two intact (N-C) assays were significantly (P < 0.01) correlated (r = 0.94), but their serum values differed by more than twofold in terms of the same BGP standard. The polyclonal assay significantly correlated with each of the intact assays (r = 0.83, 0.77), but it, too, gave different serum values for BGP. These studies demonstrate the immunochemical heterogeneity of circulating BGP, heterogeneity that is manifest even in immunoassays specific for the same region of the molecule.
我们制备了一组针对人骨钙素（BGP；骨钙蛋白）肽段的单克隆抗体，这些肽段覆盖了该分子的线性序列，具体包括BGP 1 - 12（N端）、BGP 15 - 30（中间区域）和BGP 38 - 49（C端）。在血清BGP浓度研究中，对这些抗体进行了多种两点法组合的评估。对于临床研究，我们从一组抗体中挑选出针对完整分子（N - C）的最敏感的两对抗体；我们还使用了基于BGP - C的多克隆放射免疫分析（RIA）。对于两点法，我们使用了两种吸附在聚苯乙烯珠上的N端抗体029和052，并对C端抗体663进行了放射性碘化。每种测定方法的标准品均为纯化的人BGP。通过各种测定方法测得的以下BGP血清浓度（微克/升，均值±标准误）如下：采用029 - 663测定法，正常受试者的结果为7±3，肾衰竭患者为25±8，佩吉特病患者为12±4；采用052 - 663测定法，相应结果分别为22±4、44±12和31±7；采用多克隆测定法，结果为3±0.2、13±2和5±1。两种完整分子（N - C）测定法显著相关（P < 0.01，r = 0.94），但对于相同的BGP标准品，它们的血清值相差两倍以上。多克隆测定法与每种完整分子测定法均显著相关（r = 0.83，0.77），但它给出的BGP血清值也不同。这些研究证明了循环BGP的免疫化学异质性，这种异质性甚至在针对分子同一区域的免疫测定中也有体现。
