紫外线辐照聚苯乙烯微孔板用于酶免疫测定的研究与表征

Study and Characterization of UV-irradiated Polystyrene Microtitre Plates for Use in an Enzyme Immunoassay

以重组人钙调素 (rhCaM )、亲环素 (rhCyP)、心磷脂和双链DNA (dsDNA)为包被抗原 ,建立了检测针对上述 4种抗原自身抗体的间接ELISA方法 ,并对聚苯乙烯微孔板 (PS)紫外线 (UV)辐照前后进行了对比研究 .结果发现 :PS板经UV 辐照后 ,可显著改善酶免疫分析的测定效果 ,自身抗体的测定敏感度和重复性均有显著提高 .原子力学显微镜 (AFM)表征结果则提供了改善酶免疫分析的直接证据 ,抗原分子均匀平铺于UV 辐照的PS基底表面 ,而未经辐照的PS板则抗原分子的吸附率低 ,且分布不均并有成团聚集现象 .X射线光电子能谱 (XPS)分析表明 ,PS板经UV 辐照后 ,基底表面发生了氧化并引入了含氧的活性基团 ,O/C元素比较辐照前提高了 6 9倍 ,改善了对抗原生物分子的亲水和化学反应性能 ,此亦即UV

Using recombinant human calmodulin (rhCaM), cyclophilin (rhCyP), cardiolipin and double strained DNA (dsDNA) as coating antigens, an indirect enzyme immunoassay (EIA) for autoantibodies against 4 antigens was developed. and investgated the pre- and post UV-Irradiation of polystyrene ( PS) microtitre plateas solid phase. It was found that, compared with non-treated PS, the irradiated PS plate could significantly improve the performance of assay result, and its detective sensitivity as well as reproducibility of 4 autoantibodies were significantly elevated. Surface characterizations were done by AFM, and AFM photo-pictures furnished the powerful direct evidence of improving EIA. Antigens were well-distributed on the surface of UV-irradiated PS, whereas non-treated samples shown not uniform, the percent of moleculars which adsorbed was very low and accumulative XPS analysis revealed the bottom of the wells which irradiated were oxidated, and induced peroxide reactive groups. The ratio of oxygen and carbon (O/C) was 6.9-fold higher than that of non-treated PS, and increased the surface hydrophylicity as well as chemical reactivity. This is the main reason for improved performance of UV-irradiated EIA plates.