Phase-selective synthesis of trivalent lanthanide-doped NaGdF4 nanocrystals, capped by ionic liquid cations bearing long alkyl chains, succeeded via a one-step interface-assisted ionothermal route. Owing to the existence of an interface formed between hydrophobic ionic liquids and ethylene glycol, selectively either pure cubic or hexagonal phase NaGdF4 could be obtained by changing the amount of the added surfactant, polyethyleneimine. By doping various trivalent lanthanide cations, multicolor emissions under excitation by a single wavelength could be achieved. The nanocrystals can be surface derivatized by an amphiphilic polymer and endowed with functional groups that allow the particles to not only be dispersed in aqueous solutions but also present biocompatibility. After conjugation with biotin, the nanocrystals could be used for time-resolved fluorescence resonance energy transfer biodetection.