刀额新对虾变应原的分离、鉴定与纯化

喻海琼; 刘志刚; 张帆; 胡川

doi:10.11847/zgggws2006-22-10-28

刀额新对虾变应原的分离、鉴定与纯化

Isolation, identification and purification of Metapenaeus ensis allergens

摘要

摘要:
目的通过对我国常见的刀额新对虾变应原蛋白进行分离、鉴定与纯化, 以提供虾特异性变应原用于食物变态反应疾病的诊断和治疗。
方法通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离刀额新对虾的蛋白质组分并测定其分子量, 收集过敏病人血清, 采用免疫印迹(Western-blotting)法鉴定其变应原成分, 通过离子交换层析对刀额新对虾变应原进行初步纯化, 通过疏水层析和凝胶过滤层析进一步对变应原纯化。
结果刀额新对虾有17条蛋白带, 其中主要条带有10条, 68和36 kD为可与虾过敏性病人血清IgE结合的特异性变应原; 通过各种层析方法纯化出刀额新对虾68 kD变应原, 纯度为96%。
结论对刀额新对虾变应原进行分离、鉴定和纯化, 得到高纯度的68 kD刀额新对虾变应原, 为临床虾变态反应疾病的诊断和治疗奠定基础。

Abstract:
Objective To isolate, identify and purify the allergens from shrimp(Metapenaeus ensis)and to provide specific shrimp allergens for diag nosis and immunotherapy of food allergic diseases.
Methods Proteins from Metapenaeusensis were isolated by SDS-polyacr ylamide gel electrophoresis(SDS-PAGE)and identified by Western-blotting using the sera from patients allergic to shrimp.The allergen from Metapenaeusensis was purified by Ion Ex change Chromatogr aphy, Hydrophobic Interaction Chromatography and Gel chromatography, consequently.
Results There were 17 protein bands that separated by SDS-PAG E, ten of which were major bands.The proteins whose molecular weights(MW)were 68kD and 36kD conld bind with specific IgE from sera of shrimp allergic patients, so they were specific allergens of Metapenaeusensis.The allergen with MW of 68kD was purified by serial Chromatography, with final purity of 96%.
Conclusion The allergens of Metapenaeus ensis are isolated, identified and purified.The specific allergen of Metapenaeus ensis with MW of 68kD is obtained with high purity, which may provide a basis for diagnosis and immunother apy of allergy.

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