ENRICHMENT OF AMYLOID-beta-PEPTIDES FROM CEREBROSPINAL FLUID USING SINGLE SOLID PHASE EXTRACTION BEADS AND SUBSEQUENT MASS SPECTROMETRY ANALYSIS
Bertinetti, O. Singer, K. Bertinetti, D. Hanke, S. Boerjan, Bart Reuner, K. H. Herberg, F. W.
Walter de Gruyter
Clinical Chemistry and Laboratory Medicine vol:49 pages:S772-S772
Background. Clinical diagnostics of Alzheimer‘s disease (AD) focuses on the determination of relative ratios of amyloid-β-peptides (Aβ-peptides) Aβ1-42 and Aβ1-40 in cerebrospinal fluid (CSF). Enrichment of such low abundance peptides on solid phase extraction (SPE) beads in combination with mass spectrometry (MS) provides a technique to detect biomarkers in body fluids.
Methods. Miniaturisation of the SPE technique allowed capturing and subsequent analysis of the secreted Aβ-peptides from body fluids. Salt load and high abandoned liquor proteins were depleted while simultaneously Aβ-peptides where enriched from cerebrospinal fluid on one single SPE bead. Single SPE beads were transferred to a matrix-assisted laser desorption/ionization (MALDI) target and peptides were directly eluted with 2,5-Dihydroxyacetophenone matrix and subsequently analysed on a Bruker Ultraflex MALDI-TOF/TOF with high confidence. CSF from non-demented controls spiked with different Aβ-peptides was tested against AD patient liquor.
Results. Spiked CSF samples allowed an efficient enrichment procedure with subsequent MALDI analysis within minutes. Aβ-peptide concentrations comparably with literature values (< 1µM) could be detected in sample volumes < 50µl. Peptides from patients with high unspecific protein back round (blood brain barrier defects) could still be detected. AD patient samples were tested, peak intensities/areas were determined and compared with the clinical results.
Conclusions. The single bead technique enabled a fast and sensitive analysis of Aβ-peptides in low sample volume even against high protein background.