OncoProteomics Laboratory, Department of Medical Oncology, VUmc-Cancer Center Amsterdam, VU University Medical Center, The Netherlands.
PURPOSE: For biomarker discovery in cerebrospinal fluid (CSF), removal of major serum proteins is advantageous as more CSF proteins including brain-derived proteins can be identified. Our goal was to create a reproducible discovery workflow with acceptable throughput that can identify 500-1000 CSF proteins in small volumes of CSF.
EXPERIMENTAL DESIGN: In this study, we compared the performance of two multi-affinity depletion methods in spin filter format: MARS Human 14 and Seppro-IgY-14. To this end, we analyzed depleted and bound CSF fractions isolated from 0.5 mL aliquots of the same CSF sample (n=3 per depletion method) by label-free GeLC-MS/MS-based proteomics and normalized spectral counting.
RESULTS: The whole CSF dataset contained 884 proteins identified at high confidence. Depletion spin filter performance was assessed in terms of sensitivity and reproducibility of the CSF analysis. MARS and IgY-14 spin filters yielded comparable reproducibility of protein identification (71-74%) and quantification (CV 17-18%) but a significant difference in the total number of identified CSF proteins (767 and 703 proteins, respectively).
CONCLUSIONS AND CLINICAL RELEVANCE: The MARS filter compared to IgY-14 filter provides a CSF analysis with enhanced proteome coverage. We anticipate that this enhanced sensitivity will facilitate biomarker discovery in early stages of cancer or neurological disease.Copyright © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.