Chapman M D, Heymann P W, Wilkins S R, Brown M J, Platts-Mills T A
J Allergy Clin Immunol. 1987 Aug;80(2):184-94. doi: 10.1016/0091-6749(87)90128-x.
Monoclonal, two-site radioimmunoassays (RIAs) were developed to measure allergen Der p I of Dermatophagoides pteronyssinus or Der f I of D. farinae. Microtiter plates coated with monoclonal antibody (Mab) were incubated with mite extract, and bound allergen was detected with a second 125I-labeled Mab of different epitope specificity. The Mab RIAs were very sensitive (nanogram range) and highly specific. D. pteronyssinus extracts with different concentrations of Der p I demonstrated parallel binding curves, whereas a potent D. farinae extract demonstrated less than 5% of the Der p I binding in the same assay. Similar parallel curves were obtained with several D. farinae extracts in the Der f I assay, whereas D. pteronyssinus extract demonstrated little or no binding. The Mab RIAs were compared with an inhibition RIA that measured cross-reacting determinants on both Der p I and Der f I (antigen P1 equivalent [AgP1Eq]). The results demonstrated good quantitative agreement between these assays in commercial mite and house dust extracts (mean difference 1.57 +/- 0.5-fold). Thirty house dust samples with known mite counts, Der p I, and AgP1Eq content were also compared. The summed Mab RIA values for Der p I and Der f I demonstrated a very good correlation with AgP1Eq values (r = 0.86; p less than 0.001) and with assessments of total mite-allergen content by RAST inhibition (n = 21, r = 0.77; p less than 0.001). Furthermore, in samples with more than 10 mites per 100 mg of dust, the Der p I: Der f I ratio closely correlated with the ratio of the two mites counted by microscopy (n = 15, r = 0.89; p less than 0.001). The Mab RIAs can measure allergen levels in mite or dust extracts without the need for purified allergen or affinity-purified antibodies and can readily be standardized. These assays will be useful in epidemiologic studies of allergic asthma, to assess patients' exposure to mite allergens, and the effects of avoidance regimens. Because of the long-term stability and reproducibility of the reagents, Mab-based assays for specific allergens will also play an important role in the standardization of mite and other allergen extracts.
开发了单克隆双位点放射免疫测定法（RIA）来测量粉尘螨的变应原Der p I或户尘螨的Der f I。将包被有单克隆抗体（Mab）的微量滴定板与螨提取物一起孵育，并用具有不同表位特异性的第二种125I标记的Mab检测结合的变应原。Mab RIA非常灵敏（纳克范围）且高度特异。含有不同浓度Der p I的粉尘螨提取物显示出平行的结合曲线，而一种强效的户尘螨提取物在相同测定中显示出的Der p I结合率不到5%。在Der f I测定中，几种户尘螨提取物也得到了类似的平行曲线，而粉尘螨提取物显示出很少或没有结合。将Mab RIA与一种抑制RIA进行比较，后者测量Der p I和Der f I上的交叉反应决定簇（抗原P1等效物[AgP1Eq]）。结果表明，在商业螨和屋尘提取物中，这些测定之间在定量上具有良好的一致性（平均差异1.57±0.5倍）。还比较了30个已知螨计数、Der p I和AgP1Eq含量的屋尘样本。Der p I和Der f I的Mab RIA总值与AgP1Eq值显示出非常好的相关性（r = 0.86；p<0.001），并且与RAST抑制法对总螨变应原含量的评估也具有很好的相关性（n = 21，r = 0.77；p<0.001）。此外，在每100毫克粉尘中螨数超过10只的样本中，Der p I:Der f I比值与显微镜下计数的两种螨的比值密切相关（n = 15，r = 0.89；p<0.001）。Mab RIA无需纯化的变应原或亲和纯化的抗体就能测量螨或粉尘提取物中的变应原水平，并且很容易标准化。这些测定法将有助于过敏性哮喘的流行病学研究，以评估患者接触螨变应原的情况以及避免措施的效果。由于试剂的长期稳定性和可重复性，基于Mab的特定变应原测定法在螨和其他变应原提取物的标准化方面也将发挥重要作用。
