The V-ATPase is a 14 subunit complex responsible for the acidification of intracellular vesicles and organelles. The complex is also found on the surface of osteoclasts involved in bone resorption during osteoporosis and overexpressed in some tumors. The connection of the V-ATPase with these diseases has launched a need to find inhibitors of this complex as potential therapeutics. During spermatogenesis in C.elegans the fibrous body-membranous organelles (FB-MOs) act as secretory vesicles responsible for the delivery of proteins to the surface of the sperm prior to fertilization. The FB-MOs undergo acidification in spermatids facilitated by the V-ATPase. This necessary change in pH can be indicated by the use of LysoSensor Blue (Molecular Probes) staining and the spermatids ability to mature into mature spermatozoa. Here we have established a method to screen potential inhibitors of the V-ATPase complex. Luteolin is a flavonoid compound found in vegetables, fruits, and medicinal herbs. We have examined the inhibitory effects of luteolin on the V-ATPase complex in the sperm in C.elegans. Growth assays were used to examine the toxicity of the compound. In addition to the staining for acidification of the FB-MOs, morphology of the germ cells undergoing spermatogenesis as well as DAPI staining allowed for the comparison to spe mutants. This novel approach to screening inhibitors of the V-ATPase will facilitate the screening for new therapeutic compounds.