Institute for Prevention and Occupational Medicine of the German Social Accident Insurance, Institute of the Ruhr-Universität Bochum (IPA), Bochum, Germany.
Clin Exp Allergy. 2013 Nov;43(11):1286-96. doi: 10.1111/cea.12186.
Sensitization prevalence to moulds reached from less than 10% in the general population to more than 25% in atopic and/or asthmatic subjects. To diagnose IgE-mediated mould sensitization, skin prick test (SPT) and specific IgE (sIgE) measurement are recommended. However, concordance of SPT and sIgE results is often less than 50% and standardization of the extracts is required to achieve reliable test results.
The aim of our study was to analyse mould SPT solutions (SPTs) with respect to quantity and quality of protein, antigen and human IgE-binding content as a prerequisite for further in vivo studies.
Commercial SPTs of Alternaria alternata, Aspergillus fumigatus, Cladosporium herbarum and Penicillium chrysogenum from six manufacturers as well as two in-house extracts from Aspergillus versicolor were investigated. Protein-, antigen- and IgE-binding contents were quantified by Bradford assay, sandwich ELISA and IgE-ImmunoCAP-inhibition tests. Protein composition and IgE and IgG binding were analysed by SDS-PAGE and immunoblotting, respectively.
Median protein concentrations were similar in all mould SPT extracts (90-110 μg/mL). In contrast, antigen contents and IgE-binding capacity showed a high variability with median antigen values from 4 to 118 μg/mL and IgE inhibition results between 30 to 95%. Whereas almost all SPTs of A. alternata and A. versicolor showed complete sIgE inhibition with mean values > 80%, only three extracts for A. fumigatus, two extracts for C. herbarum and none of the tested extracts for P. chrysogenum exceeded 50% sIgE reduction. Quantitative amounts of protein, antigenic and IgE-binding structures were not comparable with the quality of the corresponding protein or immunoblot pattern, with the exception of A. alternata SPTs.
Commercially available mould SPT extracts showed high variability raising the question of comparability and reliability of SPT results. Possible consequences for diagnostic test outcome will be investigated in the next step.
霉菌致敏的流行率从普通人群中的不足 10%上升到特应性和/或哮喘患者中的超过 25%。为了诊断 IgE 介导的霉菌致敏，推荐进行皮肤点刺试验（SPT）和特异性 IgE（sIgE）检测。然而，SPT 和 sIgE 结果的一致性通常低于 50%，并且需要对提取物进行标准化，以获得可靠的试验结果。
我们的研究旨在分析霉菌 SPT 溶液（SPT）的蛋白、抗原和人 IgE 结合含量的数量和质量，这是进一步进行体内研究的前提。
研究了来自 6 家制造商的Alternaria alternata、Aspergillus fumigatus、Cladosporium herbarum 和 Penicillium chrysogenum 的商业 SPT 以及来自 Aspergillus versicolor 的两种内部提取液。通过 Bradford 测定法、夹心 ELISA 和 IgE-ImmunoCAP 抑制试验定量测定蛋白、抗原和 IgE 结合含量。通过 SDS-PAGE 和免疫印迹分别分析蛋白组成和 IgE 和 IgG 结合。
所有霉菌 SPT 提取物的蛋白浓度中位数相似（90-110μg/mL）。相比之下，抗原含量和 IgE 结合能力存在很大差异，抗原值中位数范围为 4-118μg/mL，IgE 抑制结果为 30-95%。虽然 A. alternata 和 A. versicolor 的几乎所有 SPT 都表现出完全的 sIgE 抑制，平均抑制率>80%，但只有 3 种 A. fumigatus 的提取物、2 种 C. herbarum 的提取物和没有一种 P. chrysogenum 的提取物的 sIgE 抑制率超过 50%。定量的蛋白、抗原和 IgE 结合结构的数量与相应蛋白的质量或免疫印迹模式不具有可比性，除了 A. alternata 的 SPT。
市售的霉菌 SPT 提取物存在高度变异性，这引发了对 SPT 结果的可比性和可靠性的质疑。下一步将研究其对诊断试验结果的可能影响。
