Journal
ANALYST
Volume 129, Issue 9, Pages 817-822Publisher
ROYAL SOC CHEMISTRY
DOI: 10.1039/b407322c
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Funding
- NATIONAL EYE INSTITUTE [R03EY014908] Funding Source: NIH RePORTER
- NEI NIH HHS [EY014908] Funding Source: Medline
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This paper presents a highly efficient sample preparation technique for matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The purpose of the research is to use a conventional MALDI support to directly and conveniently detect sub-nM levels of peptides from volume-limited samples with physiological salt levels. In this new method, highly uniform matrix-nitrocellulose spots with a 500 mum diameter were conveniently generated by direct contact of a capillary tip to a stainless steel MALDI plate. An array of 50 microspots can be blotted from 1 muL matrix-nitrocellulose solution within 1 min. It was found that the addition of high concentration nitrocellulose to the alpha-cyano-4-hydroxycinnamic acid (CHCA) matrix solution is critical for the formation of microspots. Samples are deposited on top of those microspots and incubated for 3 min. The CHCA-nitrocellulose surface shows a significant peptide binding capability for sub-nM levels of peptide. Restricting the matrix spot diameter to 500 mum gives an analyte enrichment effect because the peptides are confined to a small solid-phase surface area. Selective peptide binding is seen even with >0.15 M salt levels. Loading small aliquots of samples with multiple applications allows low level peptide detection down to 100 pM. Push-pull perfusates collected from the rat striatum were successfully analyzed with the microspot method.
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