油菜花粉过敏原的分析、鉴定与纯化

Analysis,characterization and purification of allergen in oilseed rape pollen

目的对油菜花粉的变应原组分进行鉴定及初步的分离及纯化。方法提取油菜花粉粗提液,然后通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离油菜花粉的蛋白质组分并测定其相对分子质量,采用免疫印迹(Western blotting)法鉴定其变应原成分,并通过离子交换层析对油菜花粉变应原进行初步分离纯化,免疫印迹进行检测。结果油菜花粉粗提液有10余条蛋白带,其中相对分子质量为30 000、25 000、15 000和10 000的蛋白可与油菜花粉过敏性病人血清IgE结合,其中15 000和10 000为主要变应原;离子交换层析结果显示主要过敏原成分主要分布在Ⅰ、Ⅱ和Ⅲ峰中。结论对油菜花粉变应原进行了初步的分离、鉴定和纯化,为临床油菜花粉变态反应疾病的诊断和治疗奠定了基础。

In this paper,we aimed to characterize and purify the allergenic components of oilseed rape pollen.Firstly,rape pollen coarse extraction fluid was extracted,and the proteins and their molecular were analyzed by SDS-PAGE.The allergenic components of Oilseed rape pollen extract were separated by ion-exchange chromatography with DE-52,and then analyzed by Western-blotting.The extract of oilseed rape pollen displayed 10 protein bands,but only four bands（relative molecular weight are 30 000,25 000,15 000,and 10 000,respectively） could react with IgE in the sera of patients with oilseed rape pollen allergy.Among the four bands,two proteins were the major allergen,whose relative molecular weight was 15 000 and 10 000,and found in theⅠ,Ⅱ and,Ⅲ peaks of ion-exchange chromatography.Our study of preliminary isolation,identification and purification of the allergenic components in oilseed rape pollen will be used as a base for diagnosis and therapy oilseed rape pollen related allergy.