Antitumor immunity can be enhanced by the coordinated release and delivery of antigens and immune-stimulating agents to antigen-presenting cells via biodegradable vaccine carriers. So far, encapsulation of TLR ligands and tumor-associated antigens augmented cytotoxic T cell (CTLs) responses. Here, we compared the efficacy of the invariant NKT (iNKT) cell agonist α-galactosylceramide (α-GalCer) and TLR ligands (R848 and poly I:C) as an adjuvant for the full length ovalbumin (OVA) in PLGA nanoparticles. We observed that OVA+α-GalCer nanoparticles (NP) are superior over OVA+TLR-L NP in generating and stimulating antigen-specific cytotoxic T lymphocytes without the need for CD4(+) T cell help. Not only a 4-fold higher induction of antigen-specific T cells was observed, but also a more profound IFN-γ secretion was obtained by the addition α-GalCer. Surprisingly, we observed that mixtures of OVA containing NP with α-GalCer were ineffective, demonstrating that co-encapsulation of both α-GalCer and antigen within the same nanoparticle is essential for the observed T cell responses. Moreover, a single immunization with OVA+α-GalCer NP provided substantial protection from tumor formation and even delayed the growth of already established tumors, which coincided with a prominent and enhanced antigen-specific CD8(+) T cell infiltration. The provided evidence on the advantage of antigen and α-GalCer coencapsulation should be considered in the design of future nanoparticle vaccines for therapeutic purposes.