This paper presents a nondestructive methodology for the determination of mass and isotopic composition of large plutonium samples. The methodology is based on experimental data from measurements of gamma radiation emitted by plutonium samples under assay. Both plutonium mass and plutonium isotopic composition are derived from one experimental gamma spectrum in two energy ranges. Gamma spectrum in the middle-energy range is used for the determination of plutonium isotopic composition from experimental intensities of photo peaks belonging to different plutonium isotopes. Gamma spectrum in the high-energy range contains photo peaks of spontaneous fission products, and these data can be used for the determination of plutonium mass. The calibration curve for dependence of the count rates in photo peaks of spontaneous fission products on effective 240Pu mass was plotted based on experimental data for the enterprise-level reference plutonium samples. When processing experimental data, some corrections were introduced to account for self-absorption of gamma radiation in the plutonium samples and for neutron-induced fission reactions. The correction factors were calculated with the application of Monte Carlo methodology. The final relative errors in the determination of plutonium dioxide mass were within the range of (4 to 10)% (1) for nuclear material containers with different cooling times and different isotopic compositions of plutonium.