Vogel L, Lüttkopf D, Hatahet L, Haustein D, Vieths S
Department of Allergology, Paul-Ehrlich-Institut, Langen, Germany.
Allergy. 2005 Aug;60(8):1021-8. doi: 10.1111/j.1398-9995.2005.00803.x.
Biochemical and immunochemical methods used for batch control of allergen extracts rely on the binding of IgE molecules to allergens. They do not measure the ability of a protein to induce type I allergic reactions. Therefore, a biological assay was established that is based on the cellular mechanisms of allergies in order to assess the cross-linking capacity of allergens.
Rat basophilic leukaemia cells were transfected with cDNA coding for the human high affinity IgE receptor chains. The surface expression of the IgE-binding alpha-chain was detected by FACS analysis and the functional integration of the 'humanized' receptors into the signal transduction cascade was addressed by intracellular calcium mobilization. Mediator release was measured in response to human IgE and a variety of cross-linking allergen preparations.
Several clones were obtained that were able to bind allergen-specific human IgE. The results of the biological assay were compared with those obtained by immunochemical methods. The biological assay was used to determine the potency of allergen extracts, including highly diluted products that cannot be analysed by conventional methods.
A stable 'humanized' basophil cell line was established that will be a useful tool for the standardization and batch control of allergen extracts. Because of its high sensitivity, it can also be used to detect minute quantities of potentially allergenic proteins, e.g. in processed foods. In addition, the test may support the development of novel allergy vaccines, such as recombinant hypoallergenic molecules.
用于变应原提取物批量控制的生化和免疫化学方法依赖于IgE分子与变应原的结合。它们无法测量蛋白质诱导I型过敏反应的能力。因此，建立了一种基于过敏细胞机制的生物学检测方法，以评估变应原的交联能力。
用编码人高亲和力IgE受体链的cDNA转染大鼠嗜碱性白血病细胞。通过流式细胞术分析检测IgE结合α链的表面表达，并通过细胞内钙动员研究“人源化”受体在信号转导级联中的功能整合。测量对人IgE和各种交联变应原制剂的介质释放。
获得了几个能够结合变应原特异性人IgE的克隆。将生物学检测结果与免疫化学方法获得的结果进行比较。生物学检测用于确定变应原提取物的效价，包括无法用传统方法分析的高度稀释产品。
建立了一种稳定的“人源化”嗜碱性粒细胞系，它将成为变应原提取物标准化和批量控制的有用工具。由于其高灵敏度，它还可用于检测微量潜在变应原蛋白，例如在加工食品中。此外，该检测可能有助于新型过敏疫苗的开发，例如重组低变应原性分子。
