Invariant natural killer T (iNKT) cells are a subclass of T cells that initiates the secretion of T helper 1 and 2 cytokines after recognizing CD1d protein presented glycolipid antigens. In this Letter, we designed and synthesized a novel series of CD1d ligand α-galactosylceramides (α-GalCers) in which the acyl chain backbone of the lipid was incorporated with fluorine atoms. The <i>in vivo</i> evaluation of immunostimulatory activities revealed that the synthesized α-5-thio-galactopyranosyl-<i>N</i>-perfluorooctanoyl phytosphingosine exhibited a remarkable potency toward selectively enhancing T<sub>H</sub>1 cytokine production with the IFN γ/IL-4 ratio of 9/1, while its perfluorotetradecanoyl counterpart showed T<sub>H</sub>2 profile with an IFN γ/IL-4 ratio of 0.59/1. The analogues synthesized here would be used as probes to study lipid-protein interactions in α-GalCer/CD1d complexes.