Near-infrared (NIR) fluorescence (wavelength: 700-900 nm) is useful for in vivo imaging because of its high tissue permeability and low contamination of autofluorescence. We developed DeQODEtag system as a chemical biology tool for specific labeling of target cells using NIR fluorescence in living animals. In this system, cells expressing DeQODEtag, which is our previously obtained single-chain variable fragment (scFv) binding to a dinitrophenyl quencher, are specifically highlighted by small-molecular QODE probes consisted of a fluorophore and the quencher. We performed a chemical structure screening and obtained a high-affinity QODE probe, namely 6SiR700-pCF3oNP. A cell line with stable expression of EGFP-DeQODEtag was prepared based on a metastatic osteosarcoma cell line LM8. In a co-culture system, our probe selectively stained DeQODEtag-expressing LM8 cells against the original LM8 cells. We also prepared mouse models bearing implanted DeQODEtag-expressing LM8 cells, and performed in vivo imaging of the mice with intravenously administered QODE probes. Our developed DeQODEtag system successfully performed clear visualization of target tumor cells with NIR fluorescence in living mice. Future work will focus on applications of our system to in vivo brain Imaging.