High Throughput Intact Protein Analysis Using Infrared Matrix-Assisted Laser Desorption Electrospray Ionization Mass Spectrometry

Mass spectrometry (MS) is the primary analytical tool used to characterize proteins within the biopharmaceutical industry. Electrospray ionization (ESI) coupled to liquid chromatography is the gold standard technique for intact protein analysis. However, speed limitations prevent analysis of large sample numbers (>1000) in a day. Infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) MS, an ambient ionization MS technology, has recently been established as a platform for high throughput small molecule analysis. Here we report the application of such system for intact protein analysis. Up to 150 kDa proteins are detectable on a high throughput IR-MALDESI-MS system and we have evaluated how matrix affects signal. The system can analyze up to 22 protein samples in a second. Protein autophosphorylation, compound binding kinetics and compound modifications to a probe protein are demonstrated as applications. Top-down protein sequencing was conducted to identify a cysteine modification site. Two plate-based high throughput sample cleanup methods were coupled to IR-MALDESI-MS to enable analysis of samples containing high amounts of salts and buffers without compromising speed.

Improving sensitivity and linear dynamic range of intact protein analysis using a robust and easy to use microfluidic device

We demonstrate an integrated microfluidic LC device coupled to a QTOF capable of improving sensitivity and linearity for intact protein analysis while also tuning the charge state distributions (CSD) of whole antibodies.