化学
蛋白质聚集
粒径
纳米
纳米颗粒
流式细胞术
动态光散射
牛血清白蛋白
色谱法
纳米技术
人血清白蛋白
流动聚焦
生物物理学
化学工程
生物化学
材料科学
分子生物学
微流控
工程类
生物
物理化学
作者
Kaimin Gao,Lian Hong,Chengfeng Xue,Jing Zhou,Xiaomei Yan
标识
DOI:10.1021/acs.analchem.2c04382
摘要
Protein aggregation is one of the greatest challenges in biopharmaceuticals as it could decrease therapeutic efficacy, induce immunogenicity, and reduce shelf life of protein drugs. However, there lacks high-throughput methods than can count and size protein aggregates in the nanometer size range, especially for those smaller than 100 nm. Employing a laboratory-built nano-flow cytometer (nFCM) that enables light scattering detection of single silica nanoparticles as small as 24 nm with sizing resolution and accuracy comparable to those of electron microscopy, here, we report a new benchmark to analyze single protein aggregates as small as 40 nm. With an analysis rate of up to 10,000 particles/min, the size distribution and particle concentration of nanometer protein aggregates can be acquired in 2-3 min. Employing heat-induced aggregation of bovine serum albumin (BSA) at high concentrations as the model system, effects of different categories of excipients, including sugars, polyols, salts, and amino acids on the inhibition of protein aggregation were investigated. Strikingly enough, as high as 1010 to 1012 particles/mL of protein aggregates were observed in the size range of 40 to 200 nm for therapeutic proteins of human serum albumin injection, reconstituted recombinant human interieukin-2 solution, and human immunoglobulin injection. nFCM opens a new avenue to count and size nanometer protein aggregates, suggesting its future usability in the quality assessment and formulation promotion of therapeutic proteins.
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