Validation of Immunoassay-Based Tools for the Comprehensive Quantification of Aß40 and Aß42 Peptides in Plasma.

Recent advances in neuroimaging and cerebrospinal fluid (CSF) biomarker assays have provided evidence of a long preclinical stage of Alzheimer's disease (AD). This period is being increasingly targeted for secondary prevention trials of new therapies. In this context, the interest of a noninvasive, cost-effective amyloid-ß (Aß) blood-based test does not need to be overstated. Nevertheless, a thorough validation of these bioanalytical methods should be performed as a prerequisite for confident interpretation of clinical results. The aim of this study was to validate ELISA sandwich colorimetric ABtest40 and ABtest42 for the quantification of Aß40 and Aß42 in human plasma. The validation parameters assessed included precision, accuracy, sensitivity, specificity, recovery, and dilution linearity. ABtest40 and ABtest42 proved to be specific for their target peptide using Aß peptides with sequence similar to the target. Mean relative error in the quantification was found to be below 7.5% for both assays, with high intra-assay, inter-assay, and inter-batch precision (CV <9.0% on average). Sensitivity was assessed by determination of the limit of quantification fulfilling precision and accuracy criteria; it was established at 7.60 pg/ml and 3.60 pg/ml for ABtest40 and ABtest42, respectively. Plasma dilution linearity was demonstrated in PBS; however, dilution in a proprietary formulated buffer significantly increased the recovery of both Aß40 and Aß42 masked by matrix interactions, allowing a more comprehensive assessment of the free and total peptide levels in the plasma. In conclusion, both assays were successfully validated as tools for the quantification Aß40 and Aß42 in plasma.

Novel expression patterns of Pax3/Pax7 in early trunk neural crest and its melanocyte and non-melanocyte lineages in amniote embryos.

Neural crest cells are considered a key vertebrate feature that is studied intensively because of their relevance to development and evolution. Here we report the expression of Pax7 in the dorsal non-neural ectoderm and in the trunk neural crest of the early chick embryo. Pax7 is expressed in the trunk neural crest migrating along the ventral and dorsolateral routes. Pax7 is first downregulated in the neural crest-derived neuronal precursors, secondly in the glial, and finally in the melanocyte precursors. Conserved developmental expression in the melanocyte lineage of both Pax3 and Pax7 was evidenced in chick and quail, but only Pax3 in mouse and rat.