Quantifying pH in Malaria Using pHluorin and Flow Cytometry

Intracellular pH (pH_i) plays a critical role in the regulation of numerous biological functions where specific pH ranges are required for optimal operation within cells. Slight pH changes can impact the regulation of diverse molecular processes, including enzymatic activities, ion channels, and transporters, which all play a role in cell functions. Methods for quantifying pH_i continue to evolve and include various optical methods using fluorescent pH indicators. Here, we provide a protocol to measure pH_i in the cytosol of Plasmodium falciparum blood stage parasites by means of flow cytometry and using pHluorin2, a pH-sensitive fluorescent protein that has been introduced into the genome of the parasite.