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2,5-Dimethylfuran, known as DMF, is a promising second-generation biofuel candidate. The potential of 2,5-dimethylfuran as an additive in iso-octane (used as gasoline fuel substitute in this study) was studied. Using outwardly spherical flame method and high speed schlieren photography, laminar burning characteristics of 2,5-dimethylfuran/iso-octane (20%/80% vol., designated as D20)/air mixtures were experimentally investigated. Laminar flame speeds and Markstein lengths of D20–air mixtures were empirically organized as a function of initial pressures and temperatures. Onset of cellular structures at the flame front was observed at relatively higher initial pressures due to the combined effect of diffusional-thermal and hydrodynamic instabilities. Laminar flame speeds of D20–air mixtures are higher than those of iso-octane–air mixtures when the equivalence ratio is greater than 1.2.