MitoA is a ratiometric mass spectrometry probe that can be used for assessing changes in H2S within mitochondria in vivo. MitoA contains a triphenylphosphonium cation component that drives its accumulation in mitochondria where its aryl azide moiety selectively reacts with H2S to produce an amine product, MitoN. Quantifying the MitoN MitoA ratio by LC-MS MS reflects the mitochondrial matrix H2S concentration. In a mouse model of acute myocardial infarction with MitoA administered prior to ischemia, the MitoN MitoA ratio is increased only in the region of ischemia.