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.