Vieths S, Barber D, Chapman M, Costanzo A, Daas A, Fiebig H, Hanschmann K M, Hrabina M, Kaul S, Ledesma A, Moingeon P, Reese G, Schörner C, van Ree R, Weber B, Buchheit K H
Paul-Ehrlich-Institut, Langen, Germany.
Pharmeur Bio Sci Notes. 2012 Apr;2012:118-34.
The potency of allergen extracts is determined as total allergenic activity without consideration of their composition and the units differ from one manufacturer to another, making it very difficult to compare the different products. Recently, purified major allergens have been obtained by recombinant DNA technology and produced under Good Manufacturing Practice (GMP) conditions. In principle, such recombinant allergens could be established as reference standards and could help for the standardisation of the major allergen content of allergen extracts. Two recombinant major allergens, one from birch pollen, rBet v 1, and one from Timothy grass pollen, Phl p 5a, have been selected at the end of the CREATE programme as a potential starting point for the establishment as European Pharmacopoeia (Ph. Eur.) Reference Standards through a project run by the Biological Standardisation Programme (BSP) of the European Directorate for the Quality of Medicines & HealthCare (EDQM). To this end, bulk candidate recombinant materials, produced under GMP conditions, were procured from two European manufacturers and subsequently formulated and lyophilised. Four ELISA systems from three different manufacturers were included in the project, two for Bet v 1 and two for Phl p 5a with the aim of establishing reference methods for determination of the respective major antigens both in natural allergen extracts as well as in recombinant allergen products. The project was run in 3 phases: a preparatory and preliminary testing phase (feasibility phase or Phase 1), an extended feasibility phase carried out in 3 laboratories (Phase 2) to confirm the transferability of the methods and an international collaborative study with a large number of participating laboratories (Phase 3). This article describes the work done in Phase 1 and Phase 2, i.e. the physico-chemical and biological characterisation of the recombinant candidate reference standards, the assessment of their suitability for the intended purpose as well as the evaluation of the candidate ELISA systems. The results show that both candidate reference standards are suitable for the intended purpose. In addition, three out of the four ELISA systems that were included in the preliminary phase were found to be appropriate for further evaluation in the collaborative study which was organised in 2011. The results of the collaborative study will be published separately.
变应原提取物的效价是根据总变应原活性来确定的，而不考虑其成分，并且不同厂家的单位有所不同，这使得比较不同产品变得非常困难。最近，通过重组DNA技术获得了纯化的主要变应原，并在药品生产质量管理规范（GMP）条件下生产。原则上，此类重组变应原可被确立为参考标准，并有助于变应原提取物主要变应原含量的标准化。在CREATE项目结束时，已选择了两种重组主要变应原，一种来自桦树花粉，即rBet v 1，另一种来自梯牧草花粉，即Phl p 5a，作为通过欧洲药品与医疗保健质量管理局（EDQM）生物标准化项目（BSP）开展的一个项目确立为欧洲药典（Ph. Eur.）参考标准的潜在起点。为此，从两家欧洲制造商处采购了在GMP条件下生产的大量候选重组材料，随后进行配制和冻干。该项目纳入了来自三个不同厂家的四种酶联免疫吸附测定（ELISA）系统，其中两种用于Bet v 1，两种用于Phl p 5a，目的是建立用于测定天然变应原提取物以及重组变应原产品中各自主要抗原的参考方法。该项目分三个阶段进行：一个准备和初步测试阶段（可行性阶段或第1阶段）、在3个实验室开展的扩展可行性阶段（第2阶段）以确认方法的可转移性以及一项有大量参与实验室的国际协作研究（第3阶段）。本文描述了在第1阶段和第2阶段所做的工作，即重组候选参考标准的物理化学和生物学特性鉴定、对其是否适合预期用途的评估以及对候选ELISA系统的评价。结果表明，两种候选参考标准均适合预期用途。此外，初步阶段纳入的四种ELISA系统中有三种被认为适合在2011年组织的协作研究中作进一步评估。协作研究的结果将另行发表。
