Water is the major component of cells and tissues throughout all forms of life. 
Fluxes of water and solutes through cell membranes and epithelia are essential 
for osmoregulation and energy homeostasis. Aquaporins are membrane channels 
expressed in almost every organism and involved in the bidirectional transfer of 
water and small solutes across cell membranes. Aquaporins have important 
biological roles and have been implicated in several pathophysiological 
conditions suggesting a great translational potential in aquaporin-based 
diagnostics and therapeutics. Detecting aquaporin function is critical for 
assessing regulation and screening for new activity modulators that can prompt 
the development of efficient medicines. Appropriate methods for functional 
analysis comprising suitable cell models and techniques to accurately evaluate 
water and solute membrane permeability are essential to validate aquaporin 
function and assess short-term regulation. The present review describes 
established assays commonly used to assess aquaporin function in cells and 
tissues, as well as the experimental biophysical strategies required to reveal 
functional regulation and identify modulators, the first step for aquaporin drug 
discovery.