A novel laser desorption/ionization method using through hole porous alumina membranes

Abstract

Rationale: A novel matrix-free laser desorption/ionization method based on porous alumina membranes was developed. The porous alumina membranes have a two-dimensional (2D) ordered structure consisting of closely aligned straight through holes of sub-micron in diameter that are amenable to mass production by industrial fabrication processes.

Methods: Considering a balance between the ion generating efficiency and the mechanical strength of the membranes, the typical values for the hole diameter, open aperture ratio and membrane thickness were set to 200 nm, 50% and 5 μm, respectively. The membranes were coated with platinum on a single side that was exposed to the laser. Evaluation experiments were conducted on the feasibility of this membrane structure for an ionization method using a single peptide and mixed peptides and polyethylene glycol samples and a commercial matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometer in the positive ion mode.

Results: Results showed a softness of ionization and no sweet spot nature. The capillary action of the through holes with very high aspect ratio enables several loading protocols including sample impregnation from the surface opposite to the laser exposure side.

Conclusions: The feasibility study indicates that the through hole porous alumina membranes have several advantages in terms of usefulness over the conventional surface-assisted laser desorption ionization (SALDI) methods. The proposed novel ionization method is termed Desorption Ionization Using Through Hole Alumina Membrane (DIUTHAME).

Figure 1

An example of the through hole porous alumina membranes observed by scanning electron microscopy (SEM). The scale bar indicates 1 μm

Figure 2

Schematic view A, and photograph B, of the DIUTHAME chip in its final form

Figure 3

Protocols for loading sample solution to the DIUTHAME chip. A sample solution can be applied by either (i) loading onto the Pt coating surface of the DIUTHAME chip mounted on a target plate, or (ii) loading onto the hydrophilic surface of the DIUTHAME chip placed upside down then mounting the chip on a target plate in the right manner, or (iii) loading onto a target plate followed by mounting the DIUTHAME chip so that the droplet is covered with the effective area, depending on the solvent composition

Figure 4

Mass spectrum obtained from an aqueous solution of angiotensin II (human) by using DIUTHAME. The concentration of angiotensin II in the aqueous solution was set to 1 μM; this solution was then mixed with equal volumes of 0.2 M aqueous citric acid solution, 0.2 M aqueous diammonium hydrogen citrate solution, and acetonitrile. The final concentration of angiotensin II and water content in the solvent composition were 0.25 μM and 75%, respectively. The sample solution was loaded to the DIUTHAME chip by protocol (iii) described in the text

Figure 5

Mass spectrum obtained from an aqueous solution of the peptide mixture (Bruker Peptide Calibration Standard II) by using DIUTHAME. The concentration of the peptide contents in the aqueous solution was set to ten times higher than the standard preparation in the instructions provided by the manufacturer; this solution was then mixed with equal volumes of 0.2 M aqueous citric acid solution, 0.2 M aqueous diammonium hydrogen citrate solution. The sample solution was loaded to the DIUTHAME chip by protocol (ii) described in the text

Figure 6

Mass spectrum obtained from polyethylene glycol (PEG2000) dissolved in acetone by using DIUTHAME. The concentration of PEG2000 was 5 mg/mL. The sample solution was loaded to the DIUTHAME chip by protocol (i) described in the text