Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric Analysis of DNA on Microarrays

Abstract

Background: Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is a powerful tool in biomolecule analysis with a wide range of application possibilities, including genotyping of singlebase variations (also known as single-nucleotide polymorphisms, or SNPs) for candidate gene studies and diagnostic typing of DNA markers. We tested a method that does not require stringent purification of the nucleic acids and/or the use of modification chemistry before mass spectrometry analysis. Methods: We used an alternative direct analysis approach that allows MALDI analysis of crude DNA samples printed on microscope slides densely coated with primary amino groups that efficiently bind negatively charged DNA. After simple washing of the slides, we applied MALDI matrix and used a conventional MALDI mass spectrometer to detect DNA products. Results: We analyzed crude oligonucleotide samples and performed automated genotyping of single-base variations in 72 DNA samples. Conclusion: This procedure offers an operational short-cut compared with standard MALDI procedures for preparation of oligonucleotides, including purification, and thus is an efficient tool for genotyping applications, particularly those requiring accurate, flexible, and rapid data generation and medium throughput.

Single-nucleotide polymorphisms: analysis by mass spectrometry

“…The third approach (Fig. 1, C) uses a new microarray format that allows sample purification and detection on the same device 11 . This procedure represents an emerging technology that (still) requires specific equipment, such as nanodispensing robots.…”

Base-cleavable microarrays for the characterization of DNA and RNA oligonucleotides synthesized in situ by photolithography

“… 9 , 17 , 18 Mass spectrometry (MS) is another sensitive method which would provide final evidence of oligonucleotide identity but it has, to our knowledge, only been attempted on microarray surfaces suitable as matrices for MALDI-MS analyses. 19 – 21 …”

“…9,17,18 Mass spectrometry (MS) is another sensitive method which would provide final evidence of oligonucleotide identity but it has, to our knowledge, only been attempted on microarray surfaces suitable as matrices for MALDI-MS analyses. [19][20][21] We therefore wished to develop a method that allows for MS characterization of microarrays fabricated on standard glass microscope slides. In addition to the identification of full-length products, MS would likely detect synthetic failures, degraded material and incompletely deprotected sequences; essential information for the development of new in situ chemistries.…”

DNA Detection Using a Triple Readout Optical/AFM/MALDI Planar Microwell Plastic Chip

“…In addition, the sensitivity and robustness of DNA chips have also increased due to the use of different readout detectors, for example, electrochemical approaches, 16,17,20 light-scattering, 15,23 surface plasmon resonance, 24 atomic force microscopy, 25 and label-free methods (e.g., MALDI-MS). [26][27][28][29] The human herpes virus 5 or cytomegalovirus (CMV) has the largest genome (∼235 kb) of the human herpes virus family, which includes herpes simplex (1 and 2), Epstein-Barr, and varicella-zoster virus. 30,31 Although CMV is not highly contagious, it is the most common congenital infection (80% of the world population is infected) 32 and is particularly hazardous in immunocompromised individuals 31,32 and pregnant women (e.g., 10% of infected newborns with CMV exhibit permanent mental retardation and auditory damage).…”

“…To diagnose genetic diseases and detect infectious agents, labels such as metal nanoparticles, – enzymes, ,– and quantum dots , are usually preferred, due to their rapid and simple synthesis, reproducibility, and stability. In addition, the sensitivity and robustness of DNA chips have also increased due to the use of different readout detectors, for example, electrochemical approaches, ,, light-scattering, , surface plasmon resonance, atomic force microscopy, and label-free methods (e.g., MALDI-MS). – …”