Quantifying the sensitivity limit of ICG imaging in the presence of tissue autofluorescence

Significance: Fluorescent imaging (FI) using indocyanine green (ICG) is a powerful tool in medical diagnostics and surgery. Although numerous studies have focused on optimizing injection protocols and suppressing excitation light leakage, tissue autofluorescence has not been widely recognized as a fundamental factor limiting sensitivity. Aim: We aim to quantitatively determine the sensitivity limit for ICG detection in biological tissues, accounting for background signals from both scattered excitation light and tissue autofluorescence. Approach: using a nonfluorescent reference and a model medium with a known ICG concentration. Results: when accounting for the autofluorescence signal. Conclusions: Tissue autofluorescence is a fundamental factor limiting the sensitivity of ICG FI in the near-infrared range. The developed approach will allow for future optimization of imaging equipment and protocols for ICG and other contrast agents.