ISOELECTRIC FOCUSING PROCEDURES FOR ALLERGENIC EXTRACTS

针对过敏提取物的等电聚焦程序

• 批准号: 3804575
• 负责人: M C ANDERSON
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3804575
• 关键词: allergens; antiserum; binding proteins; cats; cross immunity; diagnosis design /evaluation; drug quality /standard; electrofocusing; human tissue; immunoglobulin E; immunological substance; western blottings

Isoelectric Focusing has proven to be a useful tool in the evaluation of allergenic extracts. This type of electrophoresis serves as a basis for examining the immunological activity of the extract, using a blotting procedure that passively transfers the focused extracts and identifies the IgE binding components in the products. The two companion methods have multiple uses. The identity of extracts submitted for standardization can be examined and compared to reference extracts with respect to IgE binding activity as well as stained IEF patterns. The optimal extraction method, in terms of IgE binding proteins generated, may be determined with these two methods. Differences in IgE binding in the sera of allergic individuals may be surveyed, and sera may be selected for testing pools with these methods, thereby assuring a fair representation of IgE binding activity in quantitative tests. A further adaptation, using select components to inhibit IgE binding in the immunoblotting, aids in the identification of cross reactive IgE binding components. Preparative IEF, using a new apparatus, has permitted the isolation of single components from allergenic extracts, separating non-IgE binding and IgE binding components in sufficient quantities for analysis and further testing. Following IEF, these components are being subjected to 2-D electrophoresis for further examination. IEF and immunoblotting has become a critical factor in the evaluation of stability studies for allergenic extracts. Using IEF as the preliminary screening method, the loss of well defined IEF banding patterns frequently indicates some loss of allergenic activity. Following evaluation of IEF, quantitative tests can be performed to confirm loss of allergenic activity. IEF has also proven to be useful in the evaluation of standardized cat extracts, with respect to demonstrating consistency between lots, and compositional differences related to allergenic source material. It has been proposed that IEF will become a release criteria for standardized cat extracts, in conjunction with Fel d I assays and protein assays that are already required.

等电聚焦已被证明是一个有用的工具，在评估 过敏性提取物。 这种类型的电泳作为基础， 使用印迹法检测提取物的免疫活性， 被动地转移集中的摘录并识别 产品中的IgE结合成分。 这两种方法有 多种用途。 提交用于标准化的浸提液的特性可 检查并与参考提取物进行IgE结合比较 活性以及染色的IEF模式。 最佳提取方法， 就产生的IgE结合蛋白而言，可以用这些来确定 两种方法 过敏性鼻炎患者血清IgE结合率的差异 可以调查个体，并且可以选择血清用于测试池 通过这些方法，从而确保IgE结合的公平代表性 在定量测试中。 进一步的适应，使用选择 在免疫印迹中抑制IgE结合的组分， 交叉反应性IgE结合组分的鉴定。 消除性IEF， 使用一种新的设备，允许分离单个组件， 从过敏原提取物中分离非IgE结合和IgE结合 足够数量的成分用于分析和进一步测试。 IEF后，这些成分将进行2-D电泳 做进一步检查 IEF和免疫印迹已经成为一个关键的 过敏性提取物稳定性研究评价中的一个因素。 使用IEF作为初步筛选方法， 带型通常表明变应原活性的某些丧失。 IEF评价后，可进行定量检测，以确认 过敏原活性丧失。 IEF也被证明是有用的， 标准化猫提取物的评价， 批次之间的一致性，以及与 过敏源材料。 有人建议，IEF将成为一个 标准化猫提取物的放行标准，与Fel d I测定和蛋白质测定已经需要。