Quantitative Multiplex Protein Assay

Custom-printed 4-plex plates were used as described previously^{20 (link)}. Plates were blocked with 150 µl 5% MSD Blocker A solution for an hour at room temperature with gentle shaking and washed 3 times with 250 µl PBS-T (0.05% tween). Peptide calibrators were diluted in MSD Diluent 35. Plates were loaded with samples and calibrators together with SULFO-TAG™ 4G8 or 6E10 detection antibody diluted in MSD Diluent 100 and incubated overnight at 4 °C with gentle shaking. After three washes with 250 µl PBS-T, 150 µl 2 × MSD read buffer was added to the wells. Plates were read by an MSD QuickPlex SQ 120 Imager and data were analyzed by MSD Workbench® software.