Detection and quantification of plasma amyloid-β by selected reaction monitoring mass spectrometry

Amyloid-beta (A beta) in human plasma was detected and quantified by an antibody-free method, selected reaction monitoring mass spectrometry (SRM-MS) in the current study. Due to its low abundance, SRM-based quantification in 10 mu L plasma was a challenge. Prior to SRM analysis, human plasma proteins as a whole were digested by trypsin and high pH reversed-phase liquid chromatography (RPLC) was used to fractionate the tryptic digests and to collect peptides, A beta(1-5), A beta(6-16), A beta(17-28) and A beta(29-40(42)) of either A beta(1-40) or A beta(1-42). Among those peptides, A beta(17-28) was selected as a surrogate to measure the total A beta level. Human plasma samples obtained from triplicate sample preparations were analyzed, obtaining 4.20 ng mL(-1) with a CV of 25.3%. Triplicate measurements for each sample preparation showed CV of <5%. Limit of quantification was obtained as 132 pM, which corresponded to 570 pg mL(-1) of A beta(1-40). Until now, most quantitative measurements of A beta in plasma or cerebrospinal fluid have required antibody-based immunoassays. Since quantification of A beta by immunoassays is highly dependent on the extent of epitope exposure due to aggregation or plasma protein binding, it is difficult to accurately measure the actual concentration of A beta in plasma. Our diagnostic method based on SRM using a surrogate peptide of A beta is promising in that actual amounts of total A beta can be measured regardless of the conformational status of the biomarker. (C) 2014 Elsevier B.V. All rights reserved.