A novel sensor fabrication concept using indicators trapped in nanoengineered ultrathin films deposited on fluorescent nanoparticles is demonstrated for oxygen sensing. The nanoscale systems are based on the quenching of tris(4,7-diphenyl-1,10-phenanthroline)ruthenium(II) immobilized within polyelectrolyte multilayers deposited on the surface of nanoparticle templates. The ionically bound nanofilms create a porous scaffold into which controlled precipitation of the chromophore is achieved using a combination of electrostatic attraction and dye insolubility in water. The fluorescent nanoparticles act as physical scaffolds and also provide a complementary spectral signature for use as an internal intensity reference. Oxygen sensors created on 100-nm yellow-green fluorescent particles exhibit a linear Stern-Volmer behavior with a quenching constant of 1.06 mM and sensitivity of 60%, which demonstrates that the dye is still partially accessible to oxygen following immobilization. To demonstrate the feasibility of intracellular metabolic monitoring with such nanoprobes, the oxygen sensors were chemically delivered into human dermal fibroblasts with no apparent loss in cell viability. The results prove that the approach to sensor production is facile and leads to sensitive systems that can be further optimized for improved response, and these findings support the further development of similar self-referenced probes toward quantitative intracellular analysis.