Rapid discovery of putative protein biomarkers of traumatic brain injury by SDS-PAGE-capillary liquid chromatography-tandem mass spectrometry.
We report the rapid discovery of putative protein biomarkers of traumatic brain injury (TBI) by SDS-PAGE-capillary liquid chromatography-tandem mass spectrometry (SDS-PAGE-Capillary LC-MS(2)). Ipsilateral hippocampus (IH) samples were collected from naive rats and rats subjected to controlled cortical impact (a rodent model of TBI). Protein database searching with 15,558 uninterpreted MS(2) spectra, collected in 3 days via data-dependent capillary LC-MS(2) of pooled cyanine dye-labeled samples separated by SDS-PAGE, identified more than 306 unique proteins. Differential proteomic analysis revealed differences in protein sequence coverage for 170 mammalian proteins (57 in naive only, 74 in injured only, and 39 of 64 in both), suggesting these are putative biomarkers of TBI. Confidence in our results was obtained by the presence of several known biomarkers of TBI (including alphaII-spectrin, brain creatine kinase, and neuron-specific enolase) in our data set. These results show that SDS-PAGE prior to in vitro proteolysis and capillary LC-MS(2) is a promising strategy for the rapid discovery of putative protein biomarkers associated with a specific physiological state (i.e., TBI) without a priori knowledge of the molecules involved.
Haskins
William E
WE
Center of Neuroproteomics and Biomarkers Research, Department of Neuroscience, University of Florida, Gainesville, Florida 32610, USA.
Kobeissy
Firas H
FH
Wolper
Regina A
RA
Ottens
Andrew K
AK
Kitlen
Jason W
JW
McClung
Scott H
SH
O'Steen
Barbara E
BE
Chow
Marjorie M
MM
Pineda
Jose A
JA
Denslow
Nancy D
ND
Hayes
Ronald L
RL
Wang
Kevin K W
KK
D000595:Amino Acid Sequence / Q000502:physiology; D000818:Animals; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D001930:Brain Injuries / Q000175:diagnosis* / Q000378:metabolism* / Q000503:physiopathology; D002853:Chromatography, Liquid / Q000379:methods; D003402:Creatine Kinase / Q000032:analysis / Q000378:metabolism; D030562:Databases, Protein; D004195:Disease Models, Animal; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006624:Hippocampus / Q000293:injuries* / Q000378:metabolism* / Q000503:physiopathology; D008297:Male; D013058:Mass Spectrometry / Q000379:methods; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000378:metabolism*; D009448:Neurochemistry / Q000295:instrumentation / Q000379:methods*; D010751:Phosphopyruvate Hydratase / Q000032:analysis / Q000378:metabolism; D011237:Predictive Value of Tests; D040901:Proteomics / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D013049:Spectrin / Q000032:analysis / Q000378:metabolism; D013997:Time Factors; D015854:Up-Regulation / Q000502:physiology
D015415:Biomarkers; D009419:Nerve Tissue Proteins; D013049:Spectrin; D003402:Creatine Kinase; D010751:Phosphopyruvate Hydratase