SARS-CoV-2 infection in hPSC-derived NPCs and 9-wk organoids. (A) Representative images of immunostaining of hNPCs with TUNEL staining, anti-Nestin, and anti–SARS-CoV-2 nucleocapsid antibody 0, 6, 12, and 48 hpi. Scale bar = 50 µm.(B) RT-PCR amplification of COVID-genome from infected hNPC cells. (C) Images of FOXG1 and PAX6 staining for dorsal neuron characterization. (D) Sample images of immunostaining of 9-wk organoids with DAPI, anti-MAP2, and anti–SARS-CoV-2 antibody 24 hpi after SARS-CoV-2 or mock infection. Note the perinuclear and neuritic staining of SARS-CoV-2 in the MAP2⁺ cell. Dashed white box corresponds to SARS-CoV-2⁺ and MAP2⁺ single-cell in 24-hpi organoid. Scale bar = 200 µm for zoomed-out images and 25 µm for zoomed-in images. (E) Sample images of immunostaining of 9-wk organoids with DAPI, anti-MAP2, or anti-Sox2 and anti–SARS-CoV-2 antibody 96 hpi after SARS-CoV-2 or mock infection. Dashed white box 1 corresponds to “1 panel” showing magnified SARS-COV-2⁺/SOX2⁻ cell in 96-hpi organoid. Dashed white box 2 corresponds to 2 panel showing SARS-COV-2⁺ and MAP2⁺ cell in 96-hpi organoid; 3 panel shows SOX2⁺/SARS-CoV-2⁺ cell in 96-hpi organoid. Scale bar = 200 µm for zoomed-out images and 25 µm for zoomed-in images. (F) Quantification of number of SARS-CoV-2⁺ cells/field of view that are either SOX2⁺ or MAP2⁺ in mock, 24-hpi, and 96-hpi organoids. (G) Organoids were stained with TUNEL to evaluate cell death 24 and 96 hpi. n = 4 organoids per condition, 12 cortical regions from two iPSC lines. Experiments were performed twice for reproducibility. Scale bar = 100 µm.