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中文摘要：
目的比较常用4种提取液提取的粉尘螨变应原致敏效果。方法分别用磷酸盐缓冲液（PBS,pH7.2）、PBST（含0.05%Tween-20的PBS）、Coca’s液、0.125mol/L NH4HCO3按料液比1︰20（W/V）的比例提取粉尘螨变应原并进行12.5%SDS-PAGE电泳,Bradford法检测提取物蛋白浓度,Western blot检测抗原性。以上述各组提取液分别致敏BALB/c小鼠,制备哮喘模型（分为PBS组,PBST组,Coca’s液、NH4HCO3组,PBS阴性组,OVA组）。采集小鼠血和支气管肺泡灌洗液（BALF）,ELISA法检测血清IgE和BALF中的IFN-γ、IL-4水平,对BALF中嗜酸性粒细胞数进行统计分析。制备肺组织病理切片,观察病理变化。结果与其他各组相比,Coca’s液提取变应原蛋白浓度最高[（274±23.6）μg/ml]（P〈0.05）;SDS-PAGE电泳检测各组提取蛋白分子质量单位均在35ku～14ku之间。ELISA检测Co-ca’s液组血清IgE含量显著高于其他组（OVA组除外）（P〈0.05）;BALF中IFN-γ、IL-4含量Coca’s液组与PBS阴性组、PBS组、PBST组和NH4HCO3组比较差异有统计学意义（P〈0.05）。结论在4种提取液中,以Coca’s液提取的粉尘螨变应原致敏效果最佳。
英文摘要：
Objective To compare the sensitizing effect of crude extracts of house dust mites obtained using 4 types of extraction solutions.Methods Crude extracts were obtained at a ratio of material：solution of 1︰20（W/V） using phosphate-buffered saline（PBS,pH7.2）,PBST（0.05% Tween-20 in PBS）,Coca＇s solution,and 0.125 mol/L NH4HCO3 solution.The crude extracts were tested with SDS-PAGE electrophoresis,detected using the Bradford method,and confirmed using Western blotting.Murine asthmatic models were prepared using the crude extracts mentioned as an allergen;mice were divided into a PBS-negative group,PBS group,PBST group,Coca＇s solution group,and NH4HCO3 group.Sera and bronchoalveolar lavage fluid（BALF） were collected from animals.The level of IgE in serum was detected with ELISA,and levels of IFN-γ and IL-4 in BALF were also determined.The number of eosinophils in BALF was also counted,and the lungs of mice were removed without lavage for histopathological examination.Results The concentration of allergen was highest when using Coca＇s solution（274±23.6 μg/ml） compared to other extracts（P0.05）.Bands were mainly located between 35 ku and 14 ku according to SDS-PAGE.The level of IgE in serum from animals sensitized with allergen extracted using Coca＇s solution was significantly higher than that in other groups（P0.05） but was not significantly different from that in the OVA group（P0.05）.The levels of IFN-γ and IL-4 in BALF followed the same pattern as IgE.Conclusion Coca＇s solution was the best solution for extracting dust mite allergens to achieve sensitization in an asthma model.
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