on 06-14-201902:36 AM - edited on 10-15-202108:39 AM by Closed Account
Oleksandr Boychenko1; Jenny Ho2; Christopher Pynn1
ASMS 2019 Poster Purpose: Whilst LC-MS has matured into a technology capable of both high proteome depth and specificity, its widespread adoption has been limited by the low-throughput and insufficient robustness of the nanoLC-MS tools typical to research. Here we demonstrate a new set of capillary-flow LC-MS (capLCMS) methods capable of large sample cohort analysis using a Thermo Scientific™ UltiMate™ 3000 RSLCnano system coupled to a Thermo Scientific™ Q Exactive™ HF-X Hybrid Quadrupole-Orbitrap™ mass spectrometer.
Methods: We optimized five low-flow LC-MS methods capable of throughputs of 180, 100, 60, 30, 24 samples per 24 hours and MS utilization from 75 to more than 90% respectively. The methods were validated using HeLa protein and crude plasma digests and showed excellent long term reproducibility and good protein coverage with more than 1000 protein groups identified with 8 min LC-MS method and 3400 protein groups within one 60 min LC-MS run.
Results: The developed low-flow LC-MS methods provide sharp and symmetric peaks that, in combination with the high sensitivity and fast acquisition speed of the Q Exactive HF-X mass spectrometer result in robust, fast and deep profiling of biological samples including cell lysate and crude plasma protein digests. Consistent results are generated over hundreds of replicate injections proving that the methods are suited to the analysis of large sample cohorts.