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3 Methods that measure the stability of biotherapeutics Ensuring that a mAb or antibody fragment therapeutic will become an effective drug requires continuous monitoring of its stability throughout the development and manufacturing processes. MAb stability is affected by many factors, including storage temperature, protein structure, concentration in solution, exposure to light or heat, and storage buffer. Some of these factors, when encountered during process development and manufacturing, can cause protein aggregation and chemical or thermal unfolding of the biotherapeutic — which can lead to impaired function and an ineffective or even harmful drug when administered to patients. Here are some of the more commonly used methods to monitor conformational and colloidal stability of a protein. Aggregation Protein aggregation affects the safety and efficacy of a biotherapeutic. Higher-order aggregates, for example, potentially cause serious immune reactions through T-cell independent pathways. Aggregation can occur throughout the mAb manufacturing process, from detection of expression in early cell culture to when the therapeutic is on the shelf, ready for administration. Scientists must use analytical methods that are highly sensitive to detecting aggregation when performing stability studies such as size-exclusion chromatography. 1 Size-exclusion chromatography (SEC) is used for the analysis and quantitation of soluble aggregates. It measures the presence of protein particles, dimers, and higher-order structures. SEC is usually performed using high-performance liquid chromatography (HPLC) or ultra- performance liquid chromatography (UPLC) with a size-exclusion column, using an absorption wavelength of 280 or 214 nm for the detection of the eluted fractions. Researchers commonly detect several peaks, with each peak representing protein particles, monomers, dimers, or higher order structures. Preferably, most mAb will be monomers as this represents a soluble and more stable protein state.

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