Stomach contents and carbon (C) and nitrogen (N) stable isotope analysis were used to evaluate trophic relationships of jumbo squid, Dosidicus gigas. Buccal masses, beaks and stomach contents of large and medium maturing-sized jumbo squid and muscle from its main prey, the myctophid Benthosema panamense, were collected in the Gulf of California, Mexico during 1996, 1997 and 1999. Both the quantified C and N-isotope ratios in muscle, and stomach content analysis revealed that larger-sized maturing squid showed a higher trophic position than medium-sized individuals. However, a discrepancy between stomach contents versus stable isotope analyses was found in evaluating trophic relationships. Simple dilution models as a function of growth were used to estimate the C and N renewal dietary shift for jumbo squid. Estimates of the initial C and N pools in D. gigas with an initial age of 70 days and 210 days indicated isotopic shifts of 32% after a threefold biomass increase and 25% after a fourfold biomass increase, respectively. Additionally, beak samples of jumbo squid were evaluated as an alternative tissue to estimate squid trophic position using stable isotopes. The results showed a significant correlation between stable isotope ratios from muscle and beak samples. Muscle isotope values were higher than beak by 1% and 4% for δ13C and δ15N respectively. A test with jumbo squid beaks collected from a stomach of a stranded sperm whale confirmed the viability of this method.