Precise Tumor Delineation via Dual Fluorescence Lifetime Imaging

Accurate delineation of tumor boundaries is critically important for tumor resection, providing support for complete tumor excision while preserving normal tissues. Fluorescence imaging, particularly in the near-infrared IIb (NIR-IIb, 1500-1700 nm) window, has garnered growing interest for tumor visualization due to its advantages, such as high sensitivity, spatial resolution, and noninvasiveness. However, tumor boundaries visualized via traditional fluorescence imaging are poorly delineated and vary with parameters such as exposure time in fluorescence intensity imaging and the background subtraction threshold in fluorescence lifetime imaging (FLIM). To address this limitation, we developed a dual-FLIM approach, wherein tumor and surrounding normal tissues were specifically stained using Er-based lanthanide-doped nanoparticles (LnNPs) with distinct fluorescence lifetimes: 0%Tm@NaYF₄-PEG-HA (tumor-targeting via hyaluronic acid, HA) and 1.5%Tm@NaYF₄-PEG, respectively. 1.5%Tm@NaYF₄-PEG exhibited a much shorter NIR-IIb fluorescence lifetime (1550 nm, corresponding to the ⁴I_13/2 → ⁴I_15/2 transition of Er³⁺) due to energy transfer between Er³⁺(⁴I_13/2) and Tm³⁺(³F₄). This dual-FLIM approach enables the clear visualization of tumor margins while eliminating boundary inconsistencies arising from subjective parameter settings. It offers a reliable solution for intraoperative margin assessment, which could enhance surgical precision and treatment decisions.