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对黑胸大蠊与美洲大蠊体部浸出液交叉抗原成分进行分析 ,为利用有关美洲大蠊分子克隆的资料进行黑胸大蠊主要变应原的克隆、测序和重组表达做前期工作。制备纯种黑胸大蠊及美洲大蠊的体部浸出液 ,采用SDS PAGE (12 %分离胶 )分离蛋白成分 ,用Coomassi亮蓝染色 ,比较两种来源蛋白成分的差异。然后将蛋白电转至PVDF膜 ,以兔抗黑胸大蠊体部浸出液 (WBE )抗体作为第一抗体 ,进行Western免疫印迹检测 ,以判断两种蟑螂的交叉抗原成分。两种蟑螂体部浸出液蛋白成分的SDS PAGE带型略有差异 ,主要是条带密度不一致 ,而从其疏密分布 (分布型式 )上看 ,两者间存在相当多的分子量相同或相近组分。经SDS PAGE分离的条带多达近 30条 ,分子量相同的组分多达 2 0余条。免疫印迹表明 ,针对兔抗黑胸大蠊体部浸出液 (WBE )抗体 (IgG )有交叉反应的抗原成分多达 10几条。黑胸大蠊与美洲大蠊体部浸出液的蛋白成分或变应原组分间存在交叉反应成分。提示 ,利用有关美洲大蠊分子克隆的资料进一步对黑胸大蠊主要变应原进行克隆、测序或重组表达等分子生物学研究具备一定的可行性。
The cross-antigen composition of leaching liquid from black-billed cockroaches and American cockroaches was analyzed in order to clone, sequence and express the main allergen of Blattella occidentalis using the information about molecular cloning of American cockroaches. Body extracts of purebred Cockroach and American cockroach were prepared and the protein fractions were separated by SDS PAGE (12% separation gel) and stained with Coomassi Brilliant Blue to compare the differences in the protein components of the two sources. Then, the protein was electrotransferred to PVDF membrane and Western blotting was performed using rabbit anti-CBT antibody (WBE) as the primary antibody to determine the cross-antigen composition of the two cockroaches. The SDS PAGE bands of the protein components of the cockroach body leachate are slightly different, mainly due to the inconsistency of band density. However, from the distribution of their density (distribution pattern), there are quite a few equivalent molecular weights or similar components . As many as 30 bands separated by SDS PAGE, as many as 20 components with the same molecular weight. Western blotting showed that as many as 10 antigens were cross-reactive against rabbit anti-CBE antibodies (IgG). There is a cross-reaction component between the protein components or allergen components of the black-throat cockroach and American cockroach body leachate. It is suggested that molecular biology research such as cloning, sequencing or recombinant expression of the main allergen of Blattella patella may be feasible by using the information about the molecular cloning of the American cockroach.