检测脂肪酸化胰岛素放射免疫分析方法的建立

目的:建立一种高灵敏度、高特异性、操作简单快捷的放射免疫分析法(RIA),用于检测比格犬血浆中的脂肪酸化胰岛素。方法:所用试剂盒主要以豚抗胰岛素抗体为主要特异性试剂,利用均相竞争抑制原理,采用平衡竞争法对比格犬血浆进行直接测定。在该系统中加入未标记的标准品或样品,则标记抗原和非标记抗原将竞争有限且定量的抗体结合位点。结果:建立了检测脂肪酸化胰岛素的RIA法并进行了确证,方法的线性范围为3.125~800μU/mL,最低检测限为3.125μU/mL,批内和批间精密度分别为4.5%~7.0%和3.9%~7.6%,冻融稳定性和稀释稳定性等良好。结论:方法学确证结果表明,本研究建立的脂肪酸化胰岛素RIA检测方法符合新生物制品临床前药代动力学研究指导原则要求,可用于脂肪酸化胰岛素在临床前药代动力学试验的检测。

Development of Radioimmunoassay Method for Quantitative Detection of Fatty Acid Acylated Insulin

Objective： To develop a specific, high sensitive and rapid radioimmunoassay（RIA） method for the quantification of fatty acid acylated insulin in beagle canine plasma. Methods： The kit mainly used the Guinea pig anti-insulin specific antibody by competitive inhibition principle, with a balanced competition method to test the canine plasma directly. If standard or sample insulin unlabeled was added to this system, there was specific immune competition response between labeled tracer and unlabeled antigen for the limited and constant. Results： An RIA was developed with a wide dynamic range of concentrations from 3.125 to 800 μU/mL. The lowest quantification of this assay was 3.125 μU/mL. Both accuracy of the intra- and inter-assay were between 4.5%-7.0% and 3.9%-7.6% respectively. Conclusion： The assay is highly sensitive, accurate, specific and reducibility over a wide dynamic range of concentrations, which was proven to be a feasible quantitative method for fatty acid aeylated insulin analysis in preelinical pharmacokinetics.