Probe-Immobilized Affinity Chromatography/Mass Spectrometry

Abstract

A new technique has been developed that provides affinity separations directly on the surface of a matrix-assisted laser desorption/ionization mass spectrometer (MALDI-MS) probe. This strategy provides both in situ identification of biomolecules through biospecific antibody/antigen interactions and molecular weight information in a rapid and facile manner. Our technique, which we call probe affinity MS, directly couples the high selectivity of immobilized affinity chromatography with the sensitivity of MALDI-MS. As this technique permits the rapid identification of antigens, ligands, and other compounds from complex biological mixtures, we believe that it will be useful in a wide range of applications in virtually all fields of the life sciences.

An immunoaffinity tandem mass spectrometry (iMALDI) assay for detection of Francisella tularensis

“…The iMALDI technique belongs to a general category of techniques which couples affinity capture with direct mass spectrometric analysis of target proteins (Experimental section and Figure 1) [32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47][48][49]. In iMALDI, anti-peptide antibodies are immobilized on sepharose beads for affinity capture, rather than on the surface of a plate, thus eliminating the need for special surface chemistry.…”

“…Unlike conventional affinity capture/elution/MALDI techniques [33,34], the "stable isotope standards with capture by anti-peptide antibodies" (SISCAPA) approach [35], or on-target elution followed by removal of affinity beads as was done by Li [50], in iMALDI the affinity beads are placed directly on the MALDI target and analyzed without prior elution of the bound analytes [32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47][48][49], which greatly reduces the sample loss and therefore increases the detection sensitivity. In iMALDI, after immuno-adsorption, the antibody beads are arranged in a microarray/spot format on the MALDI-target plate.…”

Analysis of peptides and proteins affinity-bound to iron oxide nanoparticles by MALDI MS

“…After collection, the microbeads were washed and placed on sample target, followed by analyzing with MALDI MS. Unfortunately, the presence of those particles on the sample target was reported to cause decrease in mass accuracy and resolution [25,26].Recently, nanoparticles have become interesting probes for the separation of analyte from sample solution because of their high surface area-to-volume ratio. In addition, the interference of nanoparticles during the laser desorption/ionization process may not exist because of the smallness of the particles.…”

“…After collection, the microbeads were washed and placed on sample target, followed by analyzing with MALDI MS. Unfortunately, the presence of those particles on the sample target was reported to cause decrease in mass accuracy and resolution [25,26].…”

“…The target surface was derivatized for the selective retention of analyte while removing interferences through on-target washing. Several surface derivatizations have been designed to extract and concentrate the analyte through hydrophobic interaction [10,11], ionic interaction [12,13], or immunoaffinity [14,15]. However, the sensitivity improvement was limited by the number of binding sites on the target.…”