Extended Data Fig. 1: Measurement of mitochondrial superoxide production in BMDMs using MitoNeoD and MitoSOX. | Nature Metabolism

Extended Data Fig. 1: Measurement of mitochondrial superoxide production in BMDMs using MitoNeoD and MitoSOX.

From: Pro-inflammatory macrophages produce mitochondria-derived superoxide by reverse electron transport at complex I that regulates IL-1β release during NLRP3 inflammasome activation

Extended Data Fig. 1

a-c, Mitochondrial superoxide measured with MitoNeoD in nonstimulated (NS) BMDMs untreated or treated with 10 μM MitoParaquat (MitoPQ) for 30 min or stimulated with LPS for 24 h. a, Representative images from confocal live cell imaging show MitoNeoOH, the oxidized product of MitoNeoD and Mitotracker Deep Red FM. b, Graph shows fluorescence intensity of MitoNeoOH measured as absolute intensity with Arbitrary Units (A.U.) (n = 3 (NS + MitoPQ), n = 6 (NS, 24 h LPS)). c, Quantification of MitoNeoOH fluorescence overlay with MitoTracker Deep Red in BMDMs stimulated with LPS for 24 h and NS BMDMs treated with MitoPQ (n = 3). d-f, Mitochondrial superoxide measured with MitoSOX in BMDMs stimulated with LPS for 24 h, NS or treated with 10 μM MitoParaquat for 30 min from confocal live cell imaging. e, Fluorescence intensity is measured as absolute intensity (A.U.) (n = 3). f, Quantification of MitoSOX fluorescence overlay with MitoTracker Deep Red FM in BMDMs stimulated with LPS for 24 h or NS BMDMs treated with MitoPQ (n = 3). Data are mean +/- S.E.M. of biological replicates. Scale bars on all representative live cell confocal images are 20 μm. P values displayed above graphs were calculated using two-tailed Student’s t-test for paired comparisons or one-way ANOVA for multiple comparisons.

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