Surface-active agents are organic compounds of amphiphilic nature. When the concentration of surfactants is higher than a certain value, the monomers adhere to form well-defined aggregates known as micelles. These micelles have been employed in flow injection analysis (FIA) for various purposes. In chemiluminescence-based FIA, micelles can improve sensitivity by changing the chemical structures of the reagents, facilitating intramicellar energy transfer, accelerating the reaction kinetics, or stabilizing the excited singlet states. Micelles can improve sensitivity in FIA/ultraviolet-visible by electrostatic or solubilization effects. In FIA/fluorescence, emission intensity is considerably enhanced in micellar media due to the protective environment that restricts nonemissive energy transfer processes. If FIA is coupled with flame atomic absorption, the signals are enhanced with surfactants due to the decreased surface tension that results in the generation of smaller droplets during the aspiration and nebulization processes. In addition, surfactants promote the enrichment of the analyte in the double layer at the air-water interface. The FIA/cloud point extraction technique is based on using surfactants as alternatives to organic solvents. This review discusses the different roles of micelles in FIA methods.