On-Line Bioaffinity-Electrospray Mass Spectrometry for Simultaneous Detection, Identification, and Quantification of Protein–Ligand Interactions

We describe here an on-line combination of a surface acoustic wave (SAW) biosensor with electrospray ionization mass spectrometry (SAW-ESI-MS) that enables the direct detection, identification, and quantification of affinity-bound ligands from a protein–ligand complex on a biosensor chip. A trapping column was used between the SAW-biosensor and the electrospray mass spectrometer equipped with a micro-guard column, which provides simultaneous sample concentration and desalting for the mass spectrometric analysis of the dissociated ligand. First applications of the on-line SAW-ESI-MS combination include (1), differentiation of β-amyloid (Aβ) epitope peptides bound to anti-Aβ antibodies; (2), the identification of immobilized Substance P peptide–calmodulin complex; (3), identification and quantification of the interaction of 3-nitrotyrosine-modified peptides with nitrotyrosine-specific antibodies; and (4), identification of immobilized anti-α-synuclein–human α-synuclein complex. Quantitative determinations of protein–ligand complexes by SAW yielded dissociation constants (KD) from micro-to low nanomolar sample concentrations. The on-line bioaffinity-ESI-MS combination presented here is expected to enable broad bioanalytical application to the simultaneous, label-free determination and quantification of biopolymer-ligand interactions, as diverse as antigen-antibody and lectin-carbohydrate complexes.

Graphical AbstractAn new on-line surface acoustic wave (SAW) bioaffinity system with electrospray mass spectrometry (SAW-ESI-MS) enables the simultaneous detection, identification and quantification of protein–ligand interactions.Figure optionsDownload high-quality image (69 K)Download as PowerPoint slide