Optimization and evaluation of surface-enhanced laser-desorption/ionization time-of-flight mass spectrometry for protein profiling of cerebrospinal fluid.
Cerebrospinal fluid (CSF) potentially carries an archive of peptides and small proteins relevant to pathological processes in the central nervous system (CNS) and surrounding brain tissue. Proteomics is especially well suited for the discovery of biomarkers of diagnostic potential in CSF for early diagnosis and discrimination of several neurodegenerative diseases. ProteinChip surface-enhanced laser-desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) is one such approach which offers a unique platform for high throughput profiling of peptides and small proteins in CSF. In this study, we evaluated methodologies for the retention of CSF proteins < 20 kDa in size, and identify a strategy for screening small proteins and peptides in CSF. ProteinChip array types, along with sample and binding buffer conditions, and matrices were investigated. By coupling the processing of arrays to a liquid handler reproducible and reliable profiles, with mean peak coefficients of variation < 20%, were achieved for intra- and inter-assays under selected conditions. Based on peak m/z we found a high degree of overlap between the tested array surfaces. The combination of CM10 and IMAC30 arrays was sufficient to represent between 80-90% of all assigned peaks when using either sinapinic acid or alpha-Cyano-4-hydroxycinnamic acid as the energy absorbing matrices. Moreover, arrays processed with SPA consistently showed better peak resolution and higher peak number across all surfaces within the measured mass range. We intend to use CM10 and IMAC30 arrays prepared in sinapinic acid as a fast and cost-effective approach to drive decisions on sample selection prior to more in-depth discovery of diagnostic biomarkers in CSF using alternative but complementary proteomic strategies.
Guerreiro
Nelson
N
Novartis, BioMarker Development, Exploratory Development, Klybeckstasse, CH-4002 Basel, Switzerland.
Gomez-Mancilla
Baltazar
B
Novartis, BioMarker Development, Exploratory Development, Klybeckstasse, CH-4002 Basel, Switzerland.
Charmont
Stéphane
S
Novartis, BioMarker Development, Exploratory Development, Klybeckstasse, CH-4002 Basel, Switzerland.