Numerous investigations have been devoted to visualizing latent fingerprints (LFPs) for personal identification in forensic investigation and age estimation. While simultaneous detection of chemicals in fingerprint residues has yet to be explored. Herein, fluorescent NaYF4:Yb/Tm up-conversion nanoparticles (UCNPs), which can emit blue fluorescence upon 980 nm near-infrared irradiation, have been employed in the imaging of LFPs with high contrast and high sensitivity. The furrows and ridges which provide detailed information of LFP have been displayed clearly by Tm-doped UCNPs. Meanwhile, the quantitation of nicotine residue in LFPs has been achieved based on the blue fluorescence absorbed by I2-nicotine complex, and the data was analyzed by MATLAB with modified OPTA algorithm. It was proved that the established strategy was specific, sensitive, reliable and practical toward nicotine residue in LFPs with a detection limit of 0.05 μg.