Determining Efficiency of BeatBox and Sonication Lysis Methods for Protein Extraction Using Mass Spectrometry Analysis
Introduction: Lysing of tissue is crucial for protein extraction during sample preparation for proteomic studies. Sonication is a conventional physical disruption method used during sample preparation prior to mass spectrometry analysis for proteomics studies. New tools are currently being developed to increase the efficiency of protein extraction. A new instrument developed by PreOmics™, BeatBox, uses single magnetic beads containing a proprietary coating that prevents protein binding to lyse and homogenize biological samples including tissue. Here, the lysing efficiency of sonication and BeatBox were assessed using a mass spectrometry approach. We hypothesized that tissue samples processed using BeatBox would have a significantly higher number of protein identifications compared with samples processed by sonication due to more efficient protein extraction.
Methods: Ovarian tumor tissue samples (2 samples were pooled into 1) were cryopulverized and resuspended in urea lysis buffer prior to physical lysis of tissue with sonication or BeatBox. Protein digestion was achieved using trypsin to generate tryptic peptides, and samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using a Q Exactive Plus mass spectrometer. In total, there were 3 processing replicates for sonication and BeatBox-processed samples. Proteome Discoverer 2.5 (PD) software using the SEQUEST database search algorithm was used for protein, peptide, and Peptide Spectrum Match (PSM) identification.
Results: There was no significant difference in the number of proteins (BeatBox: 1096 ± 53, Sonication: 1072 ± 30, p= 0.5685), peptides (BeatBox: 7128 ± 431, Sonication: 6866 ± 248, p= 0.4653), or PSMs (BeatBox: 10981 ± 598, Sonication: 10287 ± 359, p= 0.2074) identified between the two methods. The general trend shows an enhanced, but not significant, performance of BeatBox. Of the 983 proteins shared between BeatBox and sonication-processed samples, 348 (35.4%) had significantly higher abundances in the BeatBox-processed samples (Welch’s t-test, p < 0.05). This result indicates that tissue samples processed with BeatBox may be more efficiently lysed than sonicated samples.
Conclusion: BeatBox performs equivalently to the sonication method for total protein, peptide, and PSM count; however, it may outperform the sonication method in terms of protein quantification.