Vilja P
Department of Biomedical Sciences, University of Tampere, Finland.
J Immunol Methods. 1991 Jan 24;136(1):77-84. doi: 10.1016/0022-1759(91)90252-b.
In this study of one-step and two different two-step non-competitive avidin-biotin assays (NABAs) were developed for the measurement of a monomeric antigen (lactoferrin, LF) using polyclonal antibodies and the detection of a heterodimeric antigen (lutropin. LH) using monoclonal antibodies. The assays were based on the use of performed complexes of biotinylated antibody and avidin-peroxidase conjugate. The detection limits and intra-assay CVs of the one- and two-step NABAs were 0.1-0.5 mg/ml and 2.6-5.1% for LF, and 0.1-0.2 IU/l and 2.3-3.7% for LH, respectively. The working range was 1-100 ng/ml for the LF assay and 1-100 IU/l for the LH assay. A linear relationship with high correlation coefficients (0.979-0.992 for LF-NABAs: 0.949-0.990 for LH-NABAs) and good agreement was observed between the one- and two-step assays and the corresponding three-step NABAs used as reference methods. However, under stringent conditions the one-step assay for heterodimeric antigen was found to be sensitive to interference. The results indicate that it is possible to perform the multistep NABAs using convenient one- and two-step protocols. The one- and two-step assays also retained the advantages of the avidin-biotin system: rapidity and good sensitivity.
在本研究中，开发了一步法以及两种不同的两步法非竞争性抗生物素蛋白-生物素测定法（NABAs），分别用于使用多克隆抗体测量单体抗原（乳铁蛋白，LF）以及使用单克隆抗体检测异二聚体抗原（促黄体激素，LH）。这些测定法基于使用生物素化抗体与抗生物素蛋白-过氧化物酶共轭物的预制复合物。一步法和两步法NABAs对LF的检测限和批内变异系数分别为0.1 - 0.5 mg/ml和2.6 - 5.1%，对LH的检测限和批内变异系数分别为0.1 - 0.2 IU/l和2.3 - 3.7%。LF测定法的工作范围为1 - 100 ng/ml，LH测定法的工作范围为1 - 100 IU/l。一步法和两步法测定与用作参考方法的相应三步法NABAs之间观察到具有高相关系数的线性关系（LF - NABAs为0.979 - 0.992；LH - NABAs为0.949 - 0.990）且一致性良好。然而，在严格条件下发现一步法检测异二聚体抗原对干扰敏感。结果表明，使用便捷的一步法和两步法方案来进行多步法NABAs是可行的。一步法和两步法测定还保留了抗生物素蛋白-生物素系统的优点：快速性和良好的灵敏度。
