The liquid atmospheric pressure-matrix-assisted laser desorption/ionization (LAP-MALDI) mass spectrometry (MS) technique showed potential to enhance protein sample acquisition rates.
A new study published by the Journal of the American Society for Mass Spectrometry proposes the use of liquid atmospheric pressure-matrix-assisted laser desorption/ionization (LAP-MALDI-MS) as an alternative mass spectrometry method for ultra-high-throughput analysis of proteins, using only femtomole quantities in approximately 0.5 µL droplets (1).
The research by Bob Challen and Reiner Cramer in the Department of Chemistry at the University of Reading (Whiteknights, Reading, UK) and Michael Morris of Waters Corporation (Wilmslow, UK) sought to improve upon processes such as acoustic ejection MS (AEMS) and infrared matrix-assisted laser desorption-electrospray ionization (IR-MALDESI), which can require volumes of 20 to 50 µL for analysis.
Researchers aimed to determine if LAP-MALDI-MS could be applied to analyze intact proteins efficiently. By using femtomole quantities in 0.5 µL droplets, LAP-MALDI-MS offers an alternative to other mass spectrometry techniques that require larger volumes. The researchers used microtiter plate formats and a high-speed XY-stage actuator to achieve rapid sample acquisition rates. LAP-MALDI-MS demonstrated promising potential for improving high-throughput (HTP) protein analysis, allowing for the analysis of protein mixtures and individual proteins at low concentrations.
HTP analysis techniques have been increasingly in demand in protein therapeutics to identify post-translational modifications and the interactions of proteins with small molecules. Analyzing small- to mid-sized molecules including peptides and pharmaceuticals using HTP mass spectrometry, meanwhile, has become an extremely expeditious process using the likes of LAP-MALDI-MS, which is able to demonstrate approximately 60 samples/s.
What this study attempted to find was whether LAP-MALDI-MS, previously known to be able to efficiently handle native protein analysis, could be viable for HTP analysis of intact proteins. Some of the proteins used in this experiment were myoglobin from equine heart, cytochrome C from bovine heart, and ubiquitin from bovine erythrocytes.
The researchers first used a 96-well quarter-size microtiter plate format and then a 384-well full-size microtiter plate format. A high-speed XY-stage actuator moved the 384-well plate, resulting in sample acquisition rates of up to 10 samples/s at a data acquisition rate of 200 spectra per scan. While the sample acquisition rate was not as robust as the 60 samples/s the researchers said LAP-MALDI-MS can generate in other applications, they said there is potential for higher speeds.
Moreover, even without the SONAR acquisition mode that was deployed by the team, LAP-MALDI-MS achieved a rate of 2 samples/s. At the 10 samples/s speed, protein mixture solutions with concentrations of ≤ 2 µM could be analyzed, with individual protein solutions able to be analyzed at ≤ 0.2 µM.
In summarizing their work, the researchers concluded that LAP-MALDI-MS was both a promising and powerful tool for improving high-throughput protein analysis.
(1) Challen, B.; Morris, M.; Cramer. R. Ultra-High-Throughput and Low-Volume Analysis of Intact Proteins with LAP-MALDI MS. J. Am. Soc. Mass Spectrom. 2023. DOI: 10.1021/jasms.3c00068