Exploring the limit of detection in biomolecular interaction analysis massspectrometry (BIA/MS): detection of attomole amounts of native proteins present in complex biological mixtures

Biomolecular interaction analysis mass spectrometry (BIA/MS) offers a uniqu e combination of two highly complementary analytical techniques: biological sensing via surface plasmon resonance and matrix-assisted laser desorption /ionization time-of-flight mass spectrometry (MALDI-TOF MS), with applicati on to both functional and structural studies of native proteins. Important to the propagation of BIA/MS is the ability to detect and work with low lev els of proteins present in complex biological mixtures. In this work, the l imit of detection in BIA/MS is explored utilizing beta-2-microglobulin (bet a(2)m), an important peripheral biological marker for renal dysfunction. St andard Ppm solutions and diluted urine samples were analyzed in the surface plasmon resonance (SPR) biosensor and the binding responses measured as a function of sample concentration/dilution and flow rate through the biosens or. Following the SPR analysis of the ppm solutions that gave the lowest re liable and reproducible SPR response, the sensor surfaces were subjected to MALDI-TOF MS, yielding spectra showing selective retrieval of beta(2)m, wi th little nonspecific binding. The results indicate that attomole amounts o f beta(2)m can be captured from urine and detected reliably with SPR and MA LDI-TOF MS analysis. (C) 2000 Elsevier Science B.V. All rights reserved.