MALDI on a Chip:  Analysis of Arrays of Low-Femtomole to Subfemtomole Quantities of Synthetic Oligonucleotides and DNA Diagnostic Products Dispensed by a Piezoelectric Pipet

Abstract

A piezoelectric pipet is used to dispense arrays of low-nanoliter aliquots of matrix and DNA into individual etched wells on <1 in. silicon chips prior to their semiautomated analysis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Spectrum acquisition is expedited relative to conventional “large-spot” MALDI since the resulting miniaturized samples are approximately the same size as the irradiation area of the ionization/desorption laser; thus, searching for crystal regions from which intense analyte signals may be obtained is not necessary. Using a linear TOF instrument designed for scanning high-density arrays of samples, mass spectra from as little as 0.2 fmol (45 nM) of a 36-mer DNA have been acquired from single miniature elements. Spot-to-spot reproducibility from microdispensed samples is superior to that using conventional pipets; in less than 6 min, spectra with high signal-to-noise ratios were acquired from 100 elements containing 8 fmol of a 25-mer. Low-nanoliter quantities of DNA diagnostic products generated in primer oligo base extension reactions from PCR templates were transferred to chips and analyzed by MALDI-TOF MS, giving accurate genotyping results for single base mutations and short tandem repeat polymorphisms (microsatellites). These procedures provide enabling capabilities for extremely accurate high-throughput DNA diagnostics and sequencing based on mass spectrometry.

Spatial profiling invertebrate ganglia using MALDI MS

“…The addition of matrix has been the focus of sample preparation strategies as the liquid phase mixing of analyte and matrix is intuitively a major cause of peptide migration. Various low-volume pico-spotting and spray techniques have been developed that allow the deposition of matrix with reduced analyte migration and provide uniform matrix coverage [9,16,18,20,25,26]. In practice, however, each deposition technique possesses different characteristics that help eliminate the peptide redistribution while still providing sufficient signal and spectral resolution.…”

Quantitative mutant analysis of viral quasispecies by chip-based matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry

“…In revising our paper, we inadvertently omitted the following information. The VMH receives input from nociceptive PBsl neurons that produce the neurotransmitter CCK (23)(24)(25)(26), a peptide that suppresses food intake when injected locally to the VMH. We regret the omission of key papers that established the background for our project and apologize to the authors of those papers.…”

“…The ability to resolve oligonucleotides varying in mass by less than a single nucleotide makes MALDI-TOF mass spectrometry an excellent platform for SNP and mutant analysis. A highly automated processing platform incorporating MALDI-TOF mass spectrometry, designated DNA MassArray, has been developed (21)(22)(23). DNA MassArray incorporates regionspecific amplification of genomic DNA spanning the genetic variation, capture of one PCR product strand serving as template for a variant-specific primer extension (usually one to two bases), dispensing of nanoliter quantities of extension products onto a matrix-loaded chip array, and product analysis by MALDI-TOF mass spectrometry.…”

“…Products were washed two times with 10 mM Tris⅐HCl, pH 8.0, and denatured at 60°C for 5 min in 50 M NH 4 OH. Approximately 7 nl of the extension product was then dispensed onto a SpectroCHIP (SEQUENOM) array prespotted with a matrix of 3-hydroxypicolinic acid by using a SpectroJET (SEQUENOM) piezoelectric nanoliter pump (23). Mass spectrometric data were generated by using a modified Bruker (Billerica, MA) Biflex III MALDI-TOF mass spectrometer (SpectroREADER).…”

Microfluidic technologies for MALDI‐MS in proteomics

“…For example, Little et al employed a microfabricated target plate based on anisotropic bulk etching of silicon to form 100-element arrays of inverted pyramidal structures (800 mm square and 100 mm deep) ( Fig. 1), with each 50 nL vial serving to constrain the final target area [45]. This concept was later employed by Ekström et al [46] using 300 mm square vials with depths less than 300 mm to achieve a detection limit in the 10-50 amol range for a panel of calibration peptides.…”