Fluorescence lifetime imaging provides more possibility of in vivo multiplexing in second near infrared (NIR-II) window. However, it still faces the obstacle that fluorescent probes with differentiable lifetime often exhibit quite different fluorescence intensity, especially the short lifetime usually accompanies with a weak fluorescence intensity, resulting in the difficulty for simultaneously decoding multiplexed lifetime information due to the interference of background noise. To facilitate high-fidelity lifetime multiplexed imaging, we developed a series of Er³⁺ doped double interface fluorescent nanoprobes (Er-DINPs): α-NaYF₄@NaErF₄: Ce@NaYbF₄@NaErF₄: Ce@NaYF₄ with strong fluorescence intensity and easily distinguishable fluorescence lifetime. Both in vitro and in vivo experimental results confirmed the advantage of these probes with comparable fluorescence intensity for high-fidelity multiplexed lifetime bioimaging.

https://onlinelibrary.wiley.com/doi/10.1002/anie.202108124