CAJ | 학술논문

目的：通过改善虾类包被抗原质量，提高血清特异性IgE(sIgE)检测的敏感性。方法制备中华对虾蛋白溶液，经酶联免疫印迹实验鉴定主要的致敏蛋白组分，葡聚糖凝胶层析法初步分离大于55 ku的蛋白组分，SDS-PAGE技术分析其蛋白。分别使用虾蛋白溶液和分离的蛋白作为包被抗原包被96微孔板，采用酶斑点印迹和间接酶联免疫吸附法（间接ELISA法）初步评价纯化后的抗原是否能够提高血清sIgE检测结果的敏感性。结果酶联免疫印迹实验显示中华对虾蛋白溶液中>55 ku的蛋白组分是主要的致敏蛋白，葡聚糖凝胶层析法初步分离了>55 ku的蛋白组分，其中主要包含10条明显蛋白条带；酶斑点印迹显示，以纯化后蛋白作为包被抗原，可提高弱阳性血清斑点光密度。同时，两种包被抗原的间接ELISA结果显示，虾有效致敏成分作为包被抗原时，92.9%的虾过敏患者血清sIgE检测值升高。结论采用中华对虾有效致敏原组分作为虾过敏检测抗原能提高血清sIgE检测结果的敏感性。
목적：통과개선하류포피항원질량，제고혈청특이성IgE(sIgE)검측적민감성。방법제비중화대하단백용액，경매련면역인적실험감정주요적치민단백조분，포취당응효층석법초보분리대우55 ku적단백조분，SDS-PAGE기술분석기단백。분별사용하단백용액화분리적단백작위포피항원포피96미공판，채용매반점인적화간접매련면역흡부법（간접ELISA법）초보평개순화후적항원시부능구제고혈청sIgE검측결과적민감성。결과매련면역인적실험현시중화대하단백용액중>55 ku적단백조분시주요적치민단백，포취당응효층석법초보분리료>55 ku적단백조분，기중주요포함10조명현단백조대；매반점인적현시，이순화후단백작위포피항원，가제고약양성혈청반점광밀도。동시，량충포피항원적간접ELISA결과현시，하유효치민성분작위포피항원시，92.9%적하과민환자혈청sIgE검측치승고。결론채용중화대하유효치민원조분작위하과민검측항원능제고혈청sIgE검측결과적민감성。
Objective To improve the detecting sensitivity of serum specific IgE (sIgE) by improving the quality of coated antigen in shrimp. Methods The extracts from shrimp protein was prepared. Western blot assay was used to identify the major allergenic protein components. The protein components>55 ku were separated by Sephadex gel chromatography. SDS-PAGE technology was used to analyze proteins. Samples of shrimp protein and proteins>55 ku were used as the coat-ing antigen to coat 96 microplate respectively. Western blot assay and ELISA were used to evaluate preliminary sensitivity of the purified antigen for detecting sIgE. Results Immunoblot experiments showed that the protein>55 ku was the main aller-genic protein component of shrimp. Those >55 ku proteins were separated successfully by Sephadex gel chromatography, showing 10 identifiable bands in SDS-PAGE. Dot-pot immunoassay showed that proteins>55 ku used as coated antigens could improve the spots density of the weak serum. Meanwhile, the result of ELISA showed that sIgE detection value in-creased 92.9%in patients with shrimp allergy after coating effective antigens. Conclusion The detecting sensitivity of sIgE can be improved by using effective protein components of shrimp as coated antigens.