원문정보
초록
영어
Fast and high-throughput validation of protein products is of interest in biopharmaceutical industry. Top-down mass spectrometry (TDMS) allows for directly measuring of the intact form of proteins (i.e., proteoform) which have genetic mutations, alternative RNA splicing events and post-translational modifications. We’ve developed a new interface, called SampleStream, enabling fast and high-throughput measurement of proteoforms coupling to TDMS. The SampleStream interface consists of a fluidic channel that incorporates a molecular weight cutoff (MWCO) membrane to trap and concentrate proteins while allowing rapid buffer-exchange. The resulted proteins can be eluted from the channel then directly analyzed by high-resolution mass spectrometry. The platform offers the reduced sample processing time and quite similar sensitivity to conventional liquid chromatography (LC) technologies. We also combined immunoprecipitation with SampleStream-mass spectrometry (i.e., IP-SampleStream-MS) as a high-throughput workflow for the quantitative analysis of target proteins. We applied IP-SampleStream-MS to human serum samples to quantify proteoforms of apolipoproteins A-I (ApoA-I) and C-III (ApoC-III), which are associated with cardiometabolic charateristics such as high-density lipoprotein cholesterol (HDL-C) and obesity. We found proteoform-to-phenotype associations for ApoC-III, glycoproteoforms of which were characterized in this study.