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目的分离纯化中华绒螯蟹的主要变应原蛋白并对其进行免疫学鉴定和应用分析。方法中华绒螯蟹蛋白采用DEAE-52离子交换层析和Sephadex G-75凝胶过滤纯化,各组分用蟹过敏症患者血清以ELISA方法分析免疫反应性,活性组分用免疫印迹法测定其相对分子质量和免疫学活性。结果纯化得到两种中华绒螯蟹变应原,在ELISA检测中显示良好的免疫反应性,其相对分子质量分别为76 000和66 000。结论经离子交换层析和分子筛层析纯化的中华绒螯蟹主要变应原保持良好的免疫学活性,可作为特异性抗原应用于ELISA诊断试剂的制备。
Objective To isolate and purify the major allergenic proteins of Eriocheir sinensis and to identify and analyze the allergens. Methods Eriocheir sinensis protein was purified by DEAE-52 ion exchange chromatography and Sephadex G-75 gel filtration. Immuno-reactivity was detected by ELISA in serum of all the crab allergy sufferers. The active components were determined by immunoblotting Relative molecular mass and immunological activity. Results Two kinds of allergens of Eriocheir sinensis were purified and showed good immunoreactivity in ELISA. The relative molecular masses were 76 000 and 66 000, respectively. Conclusion The main allergens of Eriocheir sinensis purified by ion exchange chromatography and molecular sieve chromatography have good immunological activity and can be used as specific antigens in the preparation of ELISA diagnostic reagents.