生物测定
单克隆抗体
英夫利昔单抗
细胞毒性
抗体
肿瘤坏死因子α
流式细胞术
补体依赖性细胞毒性
药理学
抗体依赖性细胞介导的细胞毒性
配体结合分析
中和
免疫学
体外
化学
医学
受体
生物
生物化学
遗传学
作者
Nohemí Salinas-Jazmín,Emilio Medina‐Rivero,Marco A. Velasco-Velázquez
出处
期刊:Methods in molecular biology
日期:2021-09-04
卷期号:: 281-294
被引量:2
标识
DOI:10.1007/978-1-0716-1450-1_17
摘要
Therapeutic monoclonal antibodies (mAbs) are complex bioengineered proteins that require to be routinely characterized with robust and reliable bioassays. Infliximab was the first anti-TNFα mAb approved for use in humans and its use has revolutionized the treatment TNF-mediated inflammatory disorders. The mechanism of action (MOA) of infliximab involves its binding to soluble (s) and membrane (m) TNFα. Here, we describe two simple in vitro bioassays for the assessment of key activities of infliximab. First, a bioassay for TNFα neutralization, which evaluates the Fab binding to sTNFα and the consequent reduction in the activation of TNFα receptors and TNFα-induced expression of adhesion molecules on endothelial cells. A second bioassay evaluates the triggering of Complement-Dependent Cytotoxicity (CDC) in cells expressing mTNFα, which requires the interaction of infliximab-Fc with proteins of the complement system. In both cases, the biological responses are measured by flow cytometry, which is accessible for most laboratories. The methods reported here can be easily adapted to other therapeutic mAbs with similar MOA.
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