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After, neurons were incubated in blocking solution containing donkey anti-mouse Alexa Fluor 488 minimal cross-reactivity antibodies (AF488, 1:500 dilution, Jackson ImmunoResearch 715-545-151) and vGlut1-STAR580 nanobodies (1:500 dilution, FluoTag-X2 N1602-Ab580-L, NanoTag Biotechnologies, RRID: AB_3076007 (custom-labelling order)) for 1 hour