Rapid high mass resolution mass spectrometry using matrix-assisted ionization

Abstract

Matrix-assisted ionization (MAI) is demonstrated to be a robust and sensitive analytical method capable of analyzing proteins such as cholera toxin B-subunit and Pertussis toxin mutant from conditions containing relatively high amounts of inorganic salts, buffers, and preservatives without the need for sample clean up or concentration beforehand. By circumventing some of the sample preparation steps, MAI simplifies and speeds up the analytical workflow for biological samples in complex media. The benefits of multiply charged ions characteristic of electrospray ionization (ESI) and the robustness of matrix-assisted laser desorption/ionization (MALDI) can be obtained from a single method, making it well suited for analysis of proteins and other biomolecules at ultra-high resolution as is shown on an Orbitrap Fusion where protein subunits were resolved for which MALDI-TOF failed. MAI results are compared with those obtained with ESI, MALDI, and laserspray ionization methods and fundamental commonalities discussed.

Novel ionization processes for use in mass spectrometry: ‘Squeezing’ nonvolatile analyte ions from crystals and droplets

“…Using a temperature range from 70 to 400 °C on an Orbitrap Q‐Exactive Focus mass spectrometer produces for 3‐NBN matrix exceptionally high analyte ion abundance without the need for high voltages or a laser , while other matrices, especially typical MALDI matrices, produce less abundant analyte ions, multiply and singly charged, and only at high temperatures. The temperature can be applied directly or indirectly to the tube, typically used as an ESI inlet . Charge separation occurs after the matrix is introduced to sub‐atmospheric pressure.…”

“…Consequently, using mass spectrometers optimized for ESI‐ and MALDI‐MS, the ease of adaptation of the methods based on the new ionization processes varies. It may only require turning off the high voltages and/or laser, or it may necessitate more significant physical modification(s) to obtain analytical useful results . As an example, the currently best performing MAI matrix, 3‐NBN, on Waters SYNAPT G2 and G2S instruments works exceptionally well for analyte ionization on the intermediate pressure MALDI sources without the use of the laser, but performs best at room temperature on the ESI Z‐Spray source blocks, and above the melting point of the matrix, 114–117 °C, the analyte ion abundance essentially ceases on this instrument .…”

“…A number of different mass spectrometers, in terms of analyzer types and vendors, have been used with the novel ionization processes demonstrating the importance of pressure, temperature, and collisions in optimizing ion formation . An example is shown in Figure for bovine serum albumin (BSA), 66 kDa, using mass spectrometers to ionize this 66 kDa protein intact and multiply charged using the new ionization processes.…”

“…This matrix‐assisted ionization (MAI) method is ultimately simple and has excellent sensitivity with a limit of detection reported to be <50 attomoles and produces clean full‐scan mass spectra requiring only a few femtomoles of, for example, drugs and peptides . Mass spectra of complex mixtures can be acquired in seconds directly from biological fluids, tissue, or buffered solution, which is especially effective in conjunction with ultra‐high resolution . By placing matrix only on the specific area(s) of interest on a surface and exposing the surface to the vacuum of the mass spectrometer, ions are observed from compounds exposed to the matrix solution, allowing rapid interrogation of ‘features of interest’ .…”

“…By placing matrix only on the specific area(s) of interest on a surface and exposing the surface to the vacuum of the mass spectrometer, ions are observed from compounds exposed to the matrix solution, allowing rapid interrogation of ‘features of interest’ . The discoveries of these processes and development of suitable technologies were accomplished through fundamental research and using commercially available ion source inlets ’as is’ or with simple physical source modifications . Recently, we have begun to design and build dedicated inlets with the promise of a better understanding of the ionization processes and making improvements in applications .…”