Quantitation of amyloid beta peptides Aβ1-38, Aβ1-40, and Aβ1-42 in human cerebrospinal fluid by ultra-performance liquid chromatography-tandem mass spectrometry

Critical events in Alzheimer's disease (AD) involve an imbalance between the production and clearance of amyloid beta (A beta) peptides from the brain. Current methods for A beta quantitation rely heavily on immuno-based techniques. However, these assays require highly specific antibodies and reagents that are time-consuming and expensive to develop. Immuno-based assays are also characterized by poor dynamic ranges, cross-reactivity, matrix interferences, and dilution linearity problems. In particular, noncommercial immunoassays are especially subject to high intra- and interassay variability because they are not subject to more stringent manufacturing controls. Combinations of these factors make immunoassays more labor-intensive and often challenging to validate in support of clinical studies. Here we describe a mixed-mode solid-phase extraction method and an ultra-performance liquid chromatography tandem mass spectrometry (SPE UPLC-MS/MS) assay for the simultaneous quantitation of A beta(1-38), A beta(1-40), and A beta(1-42) from human cerebrospinal fluid (CSF). Negative ion versus positive ion species were compared using their corresponding multiple reaction monitoring (MRM) transitions, and negative ions were approximately 1.6-fold greater in intensity but lacked selectivity in matrix. The positive ion MRM assay was more than sufficient to quantify endogenous All peptides. A beta standards were prepared in artificial CSF containing 5% rat plasma, and quality control samples were prepared in three pooled CSF sources. Extraction efficiency was greater than 80% for all three peptides, and the coefficient of variation during analysis was less than 15% for all species. Mean basal levels of A beta species from three CSF pools were 1.64, 2.17, and 1.26 ng/ml for A beta(1-38); 3.24, 3.63, and 2.55 ng/ml for A beta(1-40); and 0.50, 0.63, and 0.46 ng/ml for A beta(1-42). (C) 2011 Elsevier Inc. All rights reserved.