Homogeneous time resolved fluorescence (HTRF) assay.
The HTRF assays were performed similarly to those previously described (43 ). The antibody pairs concentrations is 0.023/1.4 µg/mL (in HTRF buffer with 1% Triton X-100) were optimized for different antibody pairs and cell lines in different experiments. For human fibroblasts cells and human iPS-derived neurons, the antibody pair is 2B7/MW1, For mouse STHdh cells and mouse brain tissue, the antibody is 2B7/2166. For all the experiments, the protein concentration (by BCA, Pierce, cat. no. 23225) was measured to ensure equal loading or cell viability. Different protein concentrations or cell numbers per well were tested to ensure that the signals were in the linear range. Background corrections were performed by subtracting the background signals from blank samples.