Analysis of substance P in rat brain by means of immunoaffinity capture and matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry.
Interest in the analysis of low abundance neuropeptides particularly using matrix-assisted laser desorption/ionization time-of-flight mass-spectrometry (MALDI-TOF-MS) is increasing because these neuropeptides are essential to the mechanism of transportation and the metabolism. This article describes an immunoprecipitation procedure that is suitable for MALDI-MS analysis of substance P (SP), a neuropeptide, in rat brain tissues. Substance P was precipitated from brain tissue extracts by immunoprecipitation with antibodies directed against SP, and are analyzed by MALDI-TOF-MS. Mass spectrometric analysis showed a singly charged [M+H]+ ion peak that corresponded to the SP molecular mass and was observed with a detection error of 1.6%. The average mass errors between the observed and theoretical molecular mass were within the 0.11 Da range. Capillary zone electrophoresis analysis was subsequently performed, and the effects of the different separation parameters were examined. Beginning with milligram quantities of brain tissue, picomole quantities of SP could be detected using this method.
Suresh Babu
C V
CV
Bioanalysis and Biotransformation Research Center, Korea Institute of Science and Technology, P.O. Box 131, Cheongryang, Seoul 130-650, South Korea.
Lee
Jeongae
J
Lho
Dong Seok
DS
Yoo
Young Sook
YS
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002138:Calibration; D019075:Electrophoresis, Capillary; D047468:Immunoprecipitation; D051381:Rats; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013373:Substance P / Q000378:metabolism*
D013373:Substance P