Microscale sample deposition onto hydrophobic target plates for trace level detection of neuropeptides in brain tissue by MALDI-MS.
A sample preparation method that combines a modified target plate with a nanoscale reversed-phase column (nanocolumn) was developed for detection of neuropeptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). A gold-coated MALDI plate was modified with an octadecanethiol (ODT) self-assembled monolayer to create a hydrophobic surface that could concentrate peptide samples into a approximately 200-500-microm diameter spot. The spot sizes generated were comparable to those obtained for a substrate patterned with 200-microm hydrophilic spots on a hydrophobic substrate. The sample spots on the ODT-coated plate were 100-fold smaller than those formed on an unmodified gold plate with a 1-microl sample and generated 10 to 50 times higher mass sensitivity for peptide standards by MALDI-TOF MS. When the sample was deposited on an ODT-modified plate from a nanocolumn, the detection limit for peptides was as low as 20 pM for 5-microl samples corresponding to 80 amol deposited. This technique was used to analyze extracts of microwave-fixed tissue from rat brain striatum. Ninety-eight putative peptides were detected including several that had masses matching neuropeptides expected in this brain region such as substance P, rimorphin, and neurotensin. Twenty-three peptides had masses that matched peaks detected by capillary liquid chromatography with electrospray ionization MS.
Wei
Hui
H
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
Dean
Stacey L
SL
Parkin
Mark C
MC
Nolkrantz
Kerstin
K
O'Callaghan
James P
JP
Kennedy
Robert T
RT
D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D008297:Male; D008832:Microchemistry; D008969:Molecular Sequence Data; D009479:Neuropeptides / Q000032:analysis*; D051381:Rats; D020318:Rats, Long-Evans; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013438:Sulfhydryl Compounds / Q000737:chemistry
D009479:Neuropeptides; D013438:Sulfhydryl Compounds; C055360:n-octadecyl mercaptan