Quantitative MALDI-TOF MS of Oligonucleotides: Difference between revisions
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The advantage of MALDI MS is that it gives a sharp, predictable signal for a biomacromolecule, with great sensitivity and robustness to assay components like buffers. One disadvantage is the difficult to quantify. One approach to overcome this weakness is to use an internal standard or relative signal intensity, proper acquisition conditions, calibration plots and correction factors. | |||
The advantage of MALDI MS is that it gives a sharp, predictable signal for a biomacromolecule, with great sensitivity and robustness to assay components like buffers. One disadvantage is the difficult to quantify. One approach to overcome this weakness is to use an internal standard or relative signal intensity, proper acquisition conditions, calibration plots and correction factors. Acquisition conditions that have been found to be successful include using moderately increased laser power and at least 100 laser shots to achieve near-exhaustive ablation of a given region of the matrix, acquiring several spectra and averaging over the relative signal (analyte versus internal standard) to obtain reliable data on concentration. Some matrices give more reproducible spectra than others. For example, trihydroxyacetophenone with diammonium citrate as co-matrix fulfills these criteria and has been used to monitor selection, genotyping or footprinting assays. | |||
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''Paper on quantitative MALDI-TOF MS of oligonucleotides (cited by later papers on the topic)'' | ''Paper on quantitative MALDI-TOF MS of oligonucleotides (cited by later papers on the topic)'' | ||
D. Sarracino, C. Richert, Quantitative MALDI-TOF Spectrometry of Oligonucleotides and a Nuclease Assay. ''Bioorg. Med. Chem. Lett.,'' '''1996''', ''6'', 2543-2548. | a) D. Sarracino, C. Richert, Quantitative MALDI-TOF Spectrometry of Oligonucleotides and a Nuclease Assay. ''Bioorg. Med. Chem. Lett.,'' '''1996''', ''6'', 2543-2548. | ||
b) K. Berlin, R. K. Jain, C. Tetzlaff, C. Steinbeck, C. Richert, Spectrometrically monitored selection experiments: quantitative laser desorption mass spectrometry of small chemical libraries. ''Chem. Biol.'' '''1997''', ''4'', 63-77. | |||
c) J. Störker, J. Mayo, C. N. Tetzlaff, D. A. Sarracino, I. Schwope, C. Richert, Rapid genotyping via MALDI-monitored nuclease selection from probe libraries, ''Nature Biotechn.'' '''2000''', ''18'', 1213-1216. | |||
Revision as of 17:42, 29 June 2024
Quantitative MALDI-TOF Mass Spectrometry of Oligonucleotides
The acronym MALDI-TOF MS stands for Matrix-Assisted Laser-Desorption-Ionization Time-of-Flight Mass Spectrometry. This type of mass spectrometry was invented by Franz Hillenkamp and Michael Karas at the University of Münster.
The advantage of MALDI MS is that it gives a sharp, predictable signal for a biomacromolecule, with great sensitivity and robustness to assay components like buffers. One disadvantage is the difficult to quantify. One approach to overcome this weakness is to use an internal standard or relative signal intensity, proper acquisition conditions, calibration plots and correction factors. Acquisition conditions that have been found to be successful include using moderately increased laser power and at least 100 laser shots to achieve near-exhaustive ablation of a given region of the matrix, acquiring several spectra and averaging over the relative signal (analyte versus internal standard) to obtain reliable data on concentration. Some matrices give more reproducible spectra than others. For example, trihydroxyacetophenone with diammonium citrate as co-matrix fulfills these criteria and has been used to monitor selection, genotyping or footprinting assays.
References
MALDI Papers
a) M. Karas, F. Hillenkamp, Laser desorption ionization of proteins with molecular masses exceeding 10,000 daltons. Anal. Chem. 1988, 60, 2299–2301.
b) Berkenkamp, S.; Kirpekar, F.; Hillenkamp, F. Infrared MALDI mass spectrometry of large nucleic acids. Science 1998, 281, 260-262.
c) Kirpekar, F.; Nordhoff, E.; Larsen, L. K.; Kristiansen, K.; Roepstorff, P.; Hillenkamp, F. DNA sequence analysis by MALDI mass spectrometry. Nucleic Acids Res. 1998, 26, 2554-2559.
d) MALDI MS. A Practical Guide to Instrumentation, Methods and Applications. Edited by Franz Hillenkamp and Jasna Peter-Katalinic, Wiley-VCH Verlag, Weinheim, 2007. ISBN: 978-3-527-31440-9
Paper on quantitative MALDI-TOF MS of oligonucleotides (cited by later papers on the topic)
a) D. Sarracino, C. Richert, Quantitative MALDI-TOF Spectrometry of Oligonucleotides and a Nuclease Assay. Bioorg. Med. Chem. Lett., 1996, 6, 2543-2548.
b) K. Berlin, R. K. Jain, C. Tetzlaff, C. Steinbeck, C. Richert, Spectrometrically monitored selection experiments: quantitative laser desorption mass spectrometry of small chemical libraries. Chem. Biol. 1997, 4, 63-77.
c) J. Störker, J. Mayo, C. N. Tetzlaff, D. A. Sarracino, I. Schwope, C. Richert, Rapid genotyping via MALDI-monitored nuclease selection from probe libraries, Nature Biotechn. 2000, 18, 1213-1216.