The origin of natural gas accumulations in thermally mature basins is poorly understood. Unraveling complex contributions from hydrocarbon cracking, mixing, redox alteration, and deeper sources requires analysis of gases trapped during specific periods of basin history. Fluid inclusions can provide such samples, but their unambiguous characterization is challenging. Here we show that three-dimensional (3-D) multimodal nonlinear optical microscopy of geologic materials allows mapping and molecular identification of trapped methane and water using coherent anti-Stokes Raman scattering, imaging of crystallographic features using second harmonic generation, and identification of higher hydrocarbons using two-photon excited fluorescence. Spatially resolved, molecule-specific characterization of fluid inclusions will improve models of natural gas generation, migration, and accumulation. We believe that these broadly applicable methods will potentially transform the characterization of geological materials.