Affinity depletion versus relative protein enrichment: a side-by-side comparison of two major strategies for increasing human cerebrospinal fluid proteome coverage

AbstractCerebrospinal fluid (CSF) is in direct contact with the central nervous system. This makes human CSF an attractive source of potential biomarkers for neurologic diseases. Similarly to blood plasma, proteomic analysis of CSF is complicated by a high dynamic range of individual protein concentrations and by the presence of several highly abundant proteins. To deal with the abundant human CSF proteins, methods developed for blood plasma/serum are routinely used. Multiple affinity removal systems and protein enrichment of less abundant proteins using a combinatorial peptide ligand library are among the most frequent approaches. However, their relative impact on CSF proteome coverage has never been evaluated side-by-side in a single study. Therefore, we explored the effect of CSF depletion using MARS 14 cartridge and ProteoMiner ligand library on the number of CSF proteins identified in subsequent LC –MS/MS analysis. LC–MS/MS analysis of crude (non-treated) CSF provided roughly 500 identified proteins. Depletion of CSF by MARS 14 cartridge increased the number of identifications to nearly 800, while treatment of CSF using ProteoMiner enabled identification of 600 proteins. To explore the pot ential losses of CSF proteins during the depletion process, we also analyzed the “waste” fractions generated by both methods, i.e., proteins retained by the MARS 14 cartridge, and the molecules present in the flow-through fraction from ProteoMiner. More than 250 proteins were b...
Source: Clinical Proteomics - Category: Biochemistry Source Type: research