Boosting your biomarker and drug discovery

Introduction
Plasma membrane (PM) proteins constitute a highly diverse array of molecules embedded within the plasma membrane. They are indispensable for the proper functioning of cells by mediating interactions between the cell and its surroundings, regulating cellular processes, and synchronizing responses to extracellular signals. Their diverse functions make them essential components of cellular physiology and key targets for biomedical research and therapeutic interventions. Preparing PM proteins for LC-MS-based analysis can be challenging due to, for example, their hydrophobic nature and subcellular localization. Efficient and reliable sample preparation is critical for the success of proteomic analysis of PM proteins. BeatBox® is a fast, easy-to-use homogenizer for boosting your biomarker and drug discovery through improved identification and quantification of PM proteins from limited amounts of tissue.
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‍Material and Methods 
Input: 1-2 mg wet-weight mouse brain tissue (3 replicates).
Workflow: Tissue samples were homogenized in the BeatBox Tissue Kit 96x with iST LYSE buffer on the BeatBox for 10 min. As a control for the homogenization step, optimized bead-based ultrasonication (10 cycles, 30 sec on/off) was performed, followed by a boiling step (95 ºC, 10 min). Next, the extracted proteins were digested for 1 h and purified using the iST workflow.
LC-MS analysis: Easy nLC 1200 (ThermoFisher Scientific) coupled to timsTOF Pro (Bruker Daltonics); DDA mode; 45 min total acquisition time; data analyzed by MaxQuant software.

‍Results:
The BeatBox effectiveness was assessed by comparing the number of plasma membrane (PM) proteins identified in mouse brain samples prepared with BeatBox and a conventional bead-based sonication method. As shown in (A), the BeatBox homogenizer improved the number of identified PM proteins. Quantification of the GABAB receptor, a prominent plasma membrane protein intricately involved in neurotransmission within the central nervous system, was also performed. The samples prepared with BeatBox displayed higher protein intensities and more peptides identified of the GABAB receptor subunits, Gabbr1 and Gabbr2, compared to bead-based ultrasonication using LC-MS-based proteomic analysis.

Conclusion:
The BeatBox homogenizer outperformed conventional bead-based ultrasonication in bolstering the identification and quantification of plasma membrane proteins, making it a reliable tool in the realm of biomarker and drug discovery research.