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Development and validation of a bioassay to evaluate binding of adalimumab to cell membrane-anchored TNFα using flow cytometry detection.

08:00 EDT 30th March 2018 | BioPortfolio

Summary of "Development and validation of a bioassay to evaluate binding of adalimumab to cell membrane-anchored TNFα using flow cytometry detection."

Physicochemical and structural properties of proteins used as active pharmaceutical ingredients of biopharmaceuticals are determinant to carry out their biological activity. In this regard, the assays intended to evaluate functionality of biopharmaceuticals provide confirmatory evidence that they contain the appropriate physicochemical properties and structural conformation. The validation of the methodologies used for the assessment of critical quality attributes of biopharmaceuticals is a key requirement for manufacturing under GMP environments. Herein we present the development and validation of a flow cytometry-based methodology for the evaluation of adalimumab's affinity towards membrane-bound TNFα (mTNFα) on recombinant CHO cells. This in vitro methodology measures the interaction between an in-solution antibody and its target molecule onto the cell surface through a fluorescent signal. The characteristics evaluated during the validation exercise showed that this methodology is suitable for its intended purpose. The assay demonstrated to be accurate (r = 0.92, slope = 1.20), precise (%CV ≤ 18.31) and specific (curve fitting, r = 0.986-0.997) to evaluate binding of adalimumab to mTNFα. The results obtained here provide evidence that detection by flow cytometry is a viable alternative for bioassays used in the pharmaceutical industry. In addition, this methodology could be standardized for the evaluation of other biomolecules acting through the same mechanism of action.

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This article was published in the following journal.

Name: Journal of pharmaceutical and biomedical analysis
ISSN: 1873-264X
Pages: 235-240

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Within a eukaryotic cell, a membrane-limited body which contains chromosomes and one or more nucleoli (CELL NUCLEOLUS). The nuclear membrane consists of a double unit-type membrane which is perforated by a number of pores; the outermost membrane is continuous with the ENDOPLASMIC RETICULUM. A cell may contain more than one nucleus. (From Singleton & Sainsbury, Dictionary of Microbiology and Molecular Biology, 2d ed)

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