Instrumentation for preparative mass spectrometry

Thomas Andrew Blake, Purdue University

Abstract

Two preparative mass spectrometers have been designed and constructed as part of efforts at developing the ion soft-landing experiment into a standard proteomics tool for purifying biomolecules which retain their original bioactivity. The first preparative instrument was designed for protein microarray fabrication using the ion soft-landing technique to collect ions in defined spots on a surface after separation by mass/charge ratio. The instrument includes an atmospheric pressure ionization source, a linear ion trap (LIT) mass analyzer, an ion collection surface positioning system, and a surface loading chamber with independent vacuum pumping. Small microarrays have been prepared by isolating and soft landing individual protein or peptide ions after electrospray ionization (ESI) of mixtures. The composition and purity of the separated materials has been confirmed using independent external mass spectrometric analysis of rinse solutions of the collected spots, either by ESI-MS or by nanospray ionization (nESI)-MS. The ability to retain bioactivity after soft landing and storage on the collection surface has been demonstrated with data shown for soft-landed trypsin. Following initial characterization of the instrument, the standard tube lens/skimmer atmospheric interface was replaced with an electrodynamic ion funnel. It was found that while the ion optical device does increase ion currents up to ∼100 times, neutral contamination of the ion collection surface can occur if the ion funnel is gas-tight. The second instrument constructed was a dual-source matrix-assisted laser desorption/ionization linear ion trap electrospray ionization mass spectrometer (MALDI-LIT-ESI MS) designed to be capable of performing both a soft-landing purification and also analysis of the collected material within the same mass spectrometer. The instrument consists of a commercial MALDI-LIT MS modified by the addition of a home-built manifold housing the ion optics and electronics necessary for the addition of the ESI source. The MALDI source has been evaluated for the analysis of soft-landed biomolecules directly from the ion collection surface and has also been used to demonstrate the capability to examine the distribution of ions on the surface using MALDI-MS imaging.

Degree

Ph.D.

Advisors

Cooks, Purdue University.

Subject Area

Analytical chemistry|Biochemistry

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