Hydrogen sulfide (HS) is the most recently accepted endogenously produced gasotransmitter and is now implicated in a variety of physiological functions. In this tutorial review, our goal is to provide researchers new to the field of HS chemical biology with practical considerations, pitfalls, and best practices to enable smooth entry into investigations focused on biological HS. We present practical handling and safety considerations for working with this reactive biomolecule, and cover basic roles of HS biogenesis and action. Experimental methods for modulating HS levels, including enzymatic knockout, RNA silencing, enzymatic inhibition, and use of small molecule HS donors are highlighted. Complementing HS modulation techniques, we also highlight current strategies for HS detection and quantification.