Efficient and timely delivery of vaccine antigens to the secondary lymphoid tissue is crucial to induce protective immune responses by vaccination. However, determining the longitudinal biodistribution of injected vaccines in the body has been a challenge. Here, the near‐infrared (NIR) fluorescence imaging is reported that can efficiently enable the trafficking and biodistribution of vaccines in real time. Zwitterionic NIR fluorophores are conjugated on the surface of model vaccines and tracked the fate of bioconjugated vaccines after intradermal administration. Using an NIR fluorescence imaging system, it is possible to obtain time‐course imaging of vaccine trafficking through the lymphatics, observing notable uptake in lymph nodes with minimal nonspecific tissue interactions. Flow cytometry analysis confirmed that the uptake in lymph nodes by antigen presenting cells was highly dependent on the hydrodynamic diameter of vaccines. These results demonstrate that the combination of a real‐time NIR fluorescence imaging system and zwitterionic fluorophores is a powerful tool to determine the fate of vaccine antigens. Since such non‐specific vaccine uptake causes serious adverse reactions, this method is not only useful for optimization of vaccine design, but also for safety evaluation of clinical vaccine candidates.

Near-infrared (NIR) fluorescence imaging provides a safe and cost-efficient method for immediate data acquisition and visualization of tissues, with technical advantages including minimal autofluorescence, reduced photon absorption, and low scattering in tissue. In this review, we introduce recent advances in NIR fluorescence imaging systems for thoracic surgery that improve the identification of vital tissues and facilitate the resection of tumorous tissues. When coupled with appropriate NIR fluorophores, NIR fluorescence imaging may transform current intraoperative thoracic surgery methods by enhancing the precision of surgical procedures and augmenting postoperative outcomes through improvements in diagnostic accuracy and reductions in the remission rate.