Quaternary Ammonium and Organophosphate Impacts on Cortical Organoids

Cortical organoids were treated on alternating days between days 61 to 70 with quaternary ammonium and phosphonium compounds or organophosphate flame retardants at their approximate IC₉₀ and IC₇₅ concentrations respectively. Organoids were harvested on day 70, washed in PBS, and fixed overnight with ice-cold 4% Paraformaldehyde (Electron Microscopy Sciences, HP1–100Kit). On the following day organoids were washed with PBS and cryoprotected using a 30% sucrose solution. Organoids were then embedded in OCT and sectioned at 15 μM. Slides were washed with PBS and incubated overnight with anti-SOX10 (1:200, R&D, AF2864) and anti-APC CC1 (1:200, Millipore, MABC200), followed by labeling with Alexa Fluor-conjugated secondary antibodies (2 μg/mL, Thermo Fisher). Slides were imaged at 10x magnification using a Hamamatsu Nanozoomer S60. Quantification of positive cells was performed using QuPath software (https://qupath.github.io/)^{52 (link)}.