The feeding strategy of female yellowfin tuna (Thunnus albacares) during their reproductive cycle was investigated using a combination of different trophic tracers, i.e., stomach contents and dual stable isotope analysis, along with an assessment of ovarian development based on a histological analysis. To complete these analyses, we collected 215 female yellowfin from the Western Indian Ocean in 2009 and 2010. From these fish, we noted the ovarian development and analyzed the contents of 166 non-empty stomachs and 104 liver and muscle tissue samples. Stomach content analysis identified a large variety of prey species (45 prey families), key groups including crustaceans dominated by the swimming crab Charybdis smithii and crustacean larvae; fish dominated by the cigarfish Cubiceps pauciradiatus; and cephalopods dominated by ommastrephids Sthenoteuthis oualaniensis and Ornithoteuthis volatilis. Individuals capable of spawning appeared to feed intensively, particularly on cigarfish, during the reproductive period. From the mean reconstituted weight values of the preys, our results indicated that this intensive feeding led to increased amounts of acquired energy. The results of the stable isotope analyses, carried out on the muscle and liver tissues, indicated a clear decrease in values from north to south. These analyses also showed that liver δ15N values in spawning females were significantly lower than those in immature and developing individuals. This latter observation highlights the differences in metabolic processes that occur between tissues during ovarian development and underlines the importance of the liver in energy acquisition and mobilization in female yellowfin tuna during reproduction.