We present the synthesis, properties, and imaging applications of a new class of diazaborine-based probes (Peroxynitrite Probe-1, PNP-1) for selective peroxynitrite (ONOO-) imaging in live cells. PNP-1 features a diazaborine-based reaction motif that provides excellent discrimination between H2O2 and ONOO-, solving a persistent challenge of organoboron-based fluorescent probes for oxidative metabolite imaging. We demonstrate the utility of PNP-1 to detect endogenously produced ONOO- in live RAW 264.7 macrophages by fluorescence microscopy, with probe selectivity confirmed with inhibition of NADPH oxidases and nitric oxide synthase, the requisite enzymatic machinery for ONOO- production. Co-localization studies unexpectedly reveal preferential mitochondrial localization, which we show is dependent on the napthalimide scaffold. Taken together, our results show that diazaborines are a novel motif for selective ONOO- reactivity, positioning them for incorporation into other ONOO--specific chemical biology tools.