Protocol for assessing enzymatic and non-enzymatic hydrogen sulfide production and levels

Hydrogen sulfide (H₂S) is a key molecule in metabolism with important physiological effects. Despite its merits, detection and/or accurate determination remain challenging. Here, we present a protocol to determine H₂S levels and cumulative production using a microsulfide ion electrode and lead acetate paper. We describe steps for measurement of H₂S levels, as well as detection of enzymatic and non-enzymatic cumulative H₂S production across various biological contexts. This protocol is thus compatible with H₂S donors, tissue samples, and live cultured cells. For complete details on the use and execution of this protocol, please refer to Caliz-Molina et al.¹.