Mass Spectrometric Identification of BSA Covalently Captured onto a Chip for Atomic Force Microscopy

Abstract

Mass spectrometry (MS) is one of the main techniques for protein identification. Herein, MS has been employed for the identification of bovine serum albumin (BSA), which was covalently immobilized on the surface of a mica chip intended for investigation by atomic force microscopy (AFM). For the immobilization, two different types of crosslinkers have been used: 4-benzoylbenzoic acid N-succinimidyl ester (SuccBB) and dithiobis(succinimidyl propionate) (DSP). According to the data obtained by using an AFM-based molecular detector, the SuccBB crosslinker was more efficient in BSA immobilization than the DSP. The type of crosslinker used for protein capturing has been found to affect the results of MS identification. The results obtained herein can be applied in the development of novel systems intended for the highly sensitive analysis of proteins with molecular detectors.

MS Identification of Blood Plasma Proteins Concentrated on a Photocrosslinker-Modified Surface

“…With respect to protein concentration, the use of chemical crosslinking agents (crosslinkers) for modifying atomically smooth surfaces is also promising [ 16 , 21 , 22 ]. Based on irreversible capturing of the investigated biomolecules from the analyzed solution onto the chemically activated atomically smooth surface of the AFM chip, this approach was called chemical fishing [ 16 ]—analogous to molecular fishing [ 23 , 24 ].…”

“…Previously, optimal conditions were established for the MS identification of bovine serum albumin (BSA) captured from model protein solutions onto the surface of mica chips, modified with either succinimide or benzophenone cross-linker (DSP and SuccBB, respectively) [ 21 ]. The use of SuccBB was preferable in the case of MS identification of BSA covalently captured on the chip surface.…”