R plex human neurofilament l assay

Quantitative biochemical analyses of human Aβ soluble and insoluble fraction levels were acquired using the V-PLEX Aβ Peptide Panel 1 (6E10) (K15200G-1; Meso Scale Discovery, Rockville, MD, United States). Plates were prepared and read according to the manufacturer’s instructions. Quantitative biochemical analysis of total tau and phosphorylated tau-231 (pTau231) in soluble fractions were also obtained using the Phospho(Thr231)/Total Tau Kit (K15121D-1; Meso Scale Discovery, Rockville, MD, United States). Finally, quantitative biochemical analysis of neurofilament-light chain (NfL) in plasma was performed using the R-Plex Human Neurofilament L Assay (K1517XR-2; Meso Scale Discovery, Rockville, MD, United States).

For analysis of neurofilament light chain (NfL) in murine plasma, the R-PLEX human Neurofilament L assay (#F217X-3, MesoScale Discovery, Gaithersburg, MD, USA) employing MSD GOLD 96-well Small Spot Streptavidin plates (#L45SA-1) was used. In brief, calibrator peptide dilutions were prepared in Diluent-12, while antibody dilutions were prepared in Diluent-11 (MSD). Plasma samples were thawed on ice and measured after a 6-fold dilution with Diluent-12. For coating, plates were incubated at room temperature with continuous agitation for 60 min with 25 µL of biotinylated capture antibody diluted in Diluent-11. The plates were washed three times with wash buffer (PBS plus 0.05% Tween-20), prior to addition of 25 µL of the prepared calibrator peptide or sample dilutions per well and incubation for 60 min at room temperature with continuous shaking. After another three washing steps, 150 µL of MSD Gold Read buffer was added per well and electrochemiluminescent signals were recorded on a MESO QuickPlex SQ 120 instrument. Data were analyzed with the Discovery Workbench software package (MSD).