Technical Abstract: Surfactant phytotoxicity has been considered detrimental for enhancement of uptake, translocation, and efficacy of systemic pesticides. A technique that measured extrusion of protons by barley (Hordeum vulgare) calli into the incubation solution was developed to detect surfactant phytotoxicity. Parameters for proton extrusion by barley calli were: 0.1 mg/L 2,4-D [(2,4-dichlorophenoxy) acetic acid] in 10% (v/v) L1 medium using 125 mg barley calli in 3 ml incubation solution with continuous aeration. If foaming occurred, the continuous aeration could be replaced by rotary shaking at 100 rpm. Proton extrusion was similar for the three barley genotypes evaluated. Proton extrusion from barley calli and electrolyte leakage from potato (Solanum tuberosum) discs were compared for their ability to detect surfactant phytotoxicity. Anionic, cationic, and nonionic surfactants were evaluated for phytotoxicity. Phytotoxicity was detected at equal or lower surfactant concentrations when tested by proton extrusion (pH change) from barley calli than by electrolyte leakage (electrocunductivity) from potato tubers. Surfactant solutions with high or low pH interfered with the proton extrusion method, while surfactant solutions with high electroconductivity reduced the sensitivity of the electroconductivity method. Thus, the proton extrusion and electrolyte leakage methods compliment each other and provide more information about surfactant phytotoxicity than either method alone.