<p>The dyeability of cotton fabric with reactive dyes using a reverse micellar approach, employing saponin, a bio-based surfactant, in a non-aqueous medium of soybean oil, was investigated. Based on the best color strength (<i>K/S</i>_<i>sum</i> values) of the dyed fabrics, the dyeing parameters—surfactant-to-water mole ratio, surfactant-to-cosurfactant mole ratio, and volume of the water pool region for dye dissolution in the reverse micelles—were optimized using the one-factor-at-a-time (OFAT) approach. Experimental results revealed the optimized condition as follows: (a) 1:250 surfactant-to-water ratio; (b) 1:10 surfactant-to-cosurfactant ratio; and (c) 5% volume of the water pool for dye solubilization. The dye-encapsulated reverse micellar system was validated as a microemulsion by particle size analysis. Saponin-dyed samples exhibit better color strength, comparable color levelness, washing, and rubbing fastness compared to water-dyed samples. Scanning electron microscopic (SEM) investigation revealed no severe damage to the cotton fiber surface. The outcomes justify the applicability of saponin in non-aqueous dyeing of cotton fabric.</p>