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Background Numerous immunoassays have been developed to quantify amyloid β1-40 (Aβ40) and amyloid β1-42 (Aβ42). Nevertheless, given the low concentration of Aβ and the high levels of interfering factors in plasma, quantification of plasma Aβ is still challenging. To overcome the problems related to the specificity of Aβ immunoassays, this study aimed to develop an immunoaffinity enrichment and LC-MS/MS (IA-MS) assay. Methods We developed an IA-MS assay using antibody-labeled magnetic beads for purification and LC-MS/MS for Aβ quantification. To avoid the loss of Aβ due to aggregation in acidic buffer, we used alkaline elution buffer for immunoaffinity enrichment. The concentrations of the Aβs in plasma samples were measured, and the correlation between the plasma and cerebrospinal fluid (CSF) Aβ42/Aβ40 ratio was also evaluated. Results The intensities of the Aβ mass peaks were significantly higher with the alkaline elution buffer than with the acidic elution buffer (Aβ40: 3.6-fold, Aβ42: 5.4-fold). This assay exhibited high reproducibility (intra-assay and inter-assay precision, %CV &amp;lt;15), and the working ranges of Aβ40 and Aβ42 were determined to be 21.7 to 692.8 pg/mL and 5.6 to 180.6 pg/mL, respectively. The concentrations of Aβ40 and Aβ42 in plasma were measured by IA-MS, and the plasma Aβ42/Aβ40 ratio was correlated with the CSF Aβ42/Aβ40 ratio (rs = 0.439, P &amp;lt; 0.01). Conclusions The IA-MS assay has sufficient analytic performance for measuring endogenous Aβ40 and Aβ42 in plasma. This assay can lead to new lines of clinical discovery related to amyloid pathology.

Quantification of Amyloid-β in Plasma by Simple and Highly Sensitive Immunoaffinity Enrichment and LC-MS/MS Assay