Improving the Microdialysis Procedure for Electrospray Ionization Mass Spectrometry of Biological Samples

Abstract

Significant advances in the area of microdialysis which allowed more effective handling of small volumes (microliters) of samples, more efficient desalting and enhanced mass spectrometric detection sensitivity are described. The previously reported on‐line coupling of microdialysis with electrospray ionization (ESI) mass spectrometry has been found to be highly effective; however, direct coupling requires relatively high sample flow rates (∽2 μl min‐1) to obtain a stable ESI current compared with the flow rates of newer ESI sources (e.g. ‘microspray,’ 10–100 nl min‐1). To circumvent this major limitation imposed by the dimensions of currently available materials, the microdialysis procedure was modified to an off‐line mode in order to avoid excessive sample consumption. A more than tenfold decrease in sample consumption was achieved using the off‐line mode vs the on‐line mode, which resulted in a similar quality spectrum. In addition, several other aspects of the microdialysis approach were altered to improve its performance further: (i) an increase in dialysis temperature was found to increase the desalting efficiency greatly and therefore improve the spectrum quality; (ii) the addition of piperidine and imidazole to the dialysis buffer solution resulted in a reduction of charge states and a further increase in detection sensitivity for DNA and (iii) use of low concentrations (0–2.5 mM NH4OAc) of dialysis buffer shifted the DNA negative ions to higher charge states and produced a nearly tenfold increase in detection sensitivity and a slightly decreased desalting efficiency. Protocols for desalting different samples using microdialysis are discussed. © 1997 by John Wiley & Sons, Ltd.

Optimized suppression of adducts in polymerase chain reaction products for semi-quantitative SNP genotyping by liquid chromatography-mass spectrometry

“…Moreover, improved desalting can also be a consequence of higher temperature of the electrosprayed column effluent. This hypothesis is supported by experiments of Liu et al [37] and Walters et al [38], who reported an increased desalting performance by heating nucleic acids solutions prior to ESI-MS.…”

“…In a report by Liu et al [37], the salt tolerances of multiple buffer exchange, organic base addition, and microdialysis were studied by adding high concentrations of sodium chloride to the analyzed nucleic acid solutions. This group reported that microdialysis offered the highest desalting performance among the three techniques, which enabled the mass spectrometric detection of nucleic acids from solutions containing 0.5 mol/l sodium chloride.…”

Analysis of short tandem repeat polymorphisms by electrospray ion trap mass spectrometry

“…So far, the most successful clean-up procedures rely on ethanol precipitation or use of spin columns followed by microdialysis as a final desalting step (16,21,30,31,34,(38)(39)(40)(41)(42)(43)(44). However, these procedures are time-and labor-intensive and incur significant sample loss.…”

“…In the past, several strategies were developed to obtain sufficiently pure DNA samples. Most involved either ethanol precipitation (16,21,22,33,(38)(39)(40)(41) or centrifugation through appropriate filter systems/spin columns (30)(31)(32)34,41) followed by microdialysis (42)(43)(44) or incubation with cation exchange particles (17) for final desalting.…”

Advantages of Thermococcus kodakaraenis (KOD) DNA Polymerase for PCR-mass spectrometry based analyses

“…PCR products were ethanol precipitated [6,7] and microdialyzed [8] prior to ESI-FTICR-MS [9]. Mass spectra were acquired in the negative-ion mode using a modified ESI-FTICR mass spectrometer (IonSpec Inc., Irvine, CA) with a 9.4 tesla superconducting magnet (Cryomagnetics, Oak Ridge, TN).…”