Beatty J D, Beatty B G, Vlahos W G, Hill L R
J Immunol Methods. 1987 Jun 26;100(1-2):161-72. doi: 10.1016/0022-1759(87)90186-4.
A computerized analysis of a quantitative enzyme-linked immunoadsorbent assay (EIA) using a non-specific immunoglobulin (IgG) of known concentration as the standard has been developed for measuring specific antibody levels in serum without the need for affinity purification of the positive control antibody. The computer program utilized logit-log linear regression analysis of sigmoid serial dilution curves plus a weighted least-squares best curve fit analysis and an iterative manipulation to eliminate errant data points. The EIA was performed using serial dilutions of standard and unknown antibodies, and a double sandwich technique. A comparison of antibody levels determined by EIA using non-specific IgG as a standard relative to antibody levels determined using affinity-purified specific antibody as a standard were 1.04, 0.53, 0.48, and 0.97 for four different polyclonal antibody systems. Five monoclonal antibodies to carcinoembryonic antigen gave ratios as described above of 1.07, 1.59, 1.73, 2.32, and 2.42. The corresponding antibody affinity constants (1/mol) were 1.0 X 10(8), 3.8 X 10(8), 5.5 X 10(9), 1.8 X 10(10), and 2.6 X 10(10) respectively. This method permits accurate quantification of serum antibody levels when affinity-purified antibodies are not readily available and avoids errors due to loss of antibody activity during affinity purification.
已开发出一种计算机化分析方法，用于定量酶联免疫吸附测定（EIA），该方法使用已知浓度的非特异性免疫球蛋白（IgG）作为标准品，无需对阳性对照抗体进行亲和纯化即可测量血清中的特异性抗体水平。该计算机程序利用S形系列稀释曲线的logit-log线性回归分析、加权最小二乘法最佳曲线拟合分析以及迭代操作来消除错误数据点。EIA采用标准抗体和未知抗体的系列稀释液，并使用双夹心技术进行。对于四种不同的多克隆抗体系统，使用非特异性IgG作为标准通过EIA测定的抗体水平与使用亲和纯化的特异性抗体作为标准测定的抗体水平之比分别为1.04、0.53、0.48和0.97。五种抗癌胚抗原单克隆抗体的上述比率分别为1.07、1.59、1.73、2.32和2.42。相应的抗体亲和常数（1/mol）分别为1.0×10⁸、3.8×10⁸、5.5×10⁹、1.8×10¹⁰和2.6×10¹⁰。当不易获得亲和纯化抗体时，该方法可准确定量血清抗体水平，并避免了亲和纯化过程中抗体活性丧失导致的误差。
