Surgical microscope with integrated fluorescence lifetime imaging for 5-aminolevulinic acid fluorescence-guided neurosurgery.

SIGNIFICANCE

5-Aminolevulinic acid (5-ALA)-based fluorescence guidance in conventional neurosurgical microscopes is limited to strongly fluorescent tumor tissue. Therefore, more sensitive, intrasurgical 5-ALA fluorescence visualization is needed.

AIM

Macroscopic fluorescence lifetime imaging (FLIM) was performed ex vivo on 5-ALA-labeled human glioma tissue through a surgical microscope to evaluate its feasibility and to compare it to fluorescence intensity imaging.

APPROACH

Frequency-domain FLIM was integrated into a surgical microscope, which enabled parallel wide-field white-light and fluorescence imaging. We first characterized our system and performed imaging of two samples of suspected low-grade glioma, which were compared to histopathology.

RESULTS

Our imaging system enabled macroscopic FLIM of a 6.5  ×  6.5  mm2 field of view at spatial resolutions <20  μm. A frame of 512  ×  512  pixels with a lifetime accuracy <1  ns was obtained in 65 s. Compared to conventional fluorescence imaging, FLIM considerably highlighted areas with weak 5-ALA fluorescence, which was in good agreement with histopathology.

CONCLUSIONS

Integration of macroscopic FLIM into a surgical microscope is feasible and a promising method for improved tumor delineation.