Figure 3
From: A Simple Procedure for Creating Scalable Phenotypic Screening Assays in Human Neurons
Development and validation of HCS assay to screen for neurotoxic compounds in iNs. (A) Stepwise protocol for the 384-well plate neurotoxicity assay starting from hiPSC differentiation to HCS data analysis. (B) Representative fluorescent high content images of iNs with Channel 1 corresponding to nuclei (DAPI) and Channel 2 for neurite outgrowth (DsRed) and a composite merged image of the 2 channels. The groups selected corresponded to low (DMSO treated) and high control (10 µM Brefeldin A-BFA treated) wells. (C) Representative fluorescent high content image processing of same images where neurites were traced according to predefined optimized parameters under the neuronal profiling module of the CellInsight software. Each image corresponds to 1 field of view and each well has 4 fields of views using the 5X objective. (D) Automated quantification of valid nucleus count, neurite count, neurite length from iNs treated with control compounds DMSO and 10 µM BFA. Data are from 4 independent experiments, total n = 12 biological replicates per experiment; values are means ± stdev. **p < 0.0001.
