Real-Time Visualization of Tissue Surface Biochemical Features Derived From Fluorescence Lifetime Measurements

Abstract

Fiber based fluorescence lifetime imaging has shown great potential for intraoperative diagnosis and guidance of surgical procedures. Here we describe a novel method addressing a significant challenge for the practical implementation of this technique, i.e. the real-time display of the quantified biochemical or functional tissue properties superimposed on the interrogated area. Specifically, an aiming beam (450 nm) generated by a continuous-wave laser beam was merged with the pulsed fluorescence excitation light in a single delivery/collection fiber and then imaged and segmented using a color-based algorithm. We demonstrate that this approach enables continuous delineation of the interrogated location and dynamic augmentation of the acquired frames with the corresponding fluorescence decay parameters. The method was evaluated on a fluorescence phantom and fresh tissue samples. Current results demonstrate that 34 frames per second can be achieved for augmenting videos of 640×512 pixels resolution. Also we show that the spatial resolution of the fluorescence lifetime map depends on the tissue optical properties, the scanning speed, and the frame rate. The dice similarity coefficient between the fluorescence phantom and the reconstructed maps was estimated to be as high as 93%. The reported method could become a valuable tool for augmenting the surgeon’s field of view with diagnostic information derived from the analysis of fluorescence lifetime data in real-time using handheld, automated, or endoscopic scanning systems. Current method provides also a means for maintaining the tissue light exposure within safety limits. This study provides a framework for using an aiming beam with other point spectroscopy applications.

Real‐time fiber‐based fluorescence lifetime imaging with synchronous external illumination: A new path for clinical translation

“…In contrast, a single point instrument lacks the spatial resolution provided by an imaging platform, which is essential in many clinical applications requiring fine margin assessment such as identification of tumor margins during resection surgery. In order to address this limitation, we followed an approach that is conceptually similar to that previously published by Gorpas et al It comprises two key elements: (a) a CW laser source that is added to the optical setup and superimposed with the excitation beam at the sample end to provide a visual reference that guides the operator during measurements; (b) a color camera is added to the setup to image the sample from a fixed position. The camera allows recording the location of fluorescence measurements as the operator moves the fiber through different regions in the sample.…”

“…The sensitivity of TCSPC to background light is perhaps the greatest hindrance to the adoption of this technique in medical procedures, since turning off all sources of background light is either impossible or, in the limit, a major disruption to the clinical workflow and thus a potential hazard. We note that other fluorescence lifetime measurement techniques, such as pulse sampling, are suitable for real‐time measurements under bright background, since the detection technique is analog and the low frequency signal of background room light can be filtered accordingly . However, the pulse sampling technique has lower sensitivity and dynamic range compared to TCSPC and thus provides lower specificity in the autofluorescence readout.…”

Rapid fluorescence lifetime estimation with modified phasor approach and Laguerre deconvolution: a comparative study

“…Our previous work has explored the application of fluorescence lifetime imaging (FLIm) for oral carcinoma detection [9], atherosclerotic plaque assessment [10] and characterization of engineered tissues [11]. More recently we have also reported the ability to measure lifetime from tissue and to augment white-light video frames in real-time [12, 13]. Quantification of emission spectrum and lifetime is performed through different analysis tools.…”

Label-free optical imaging technologies for rapid translation and use during intraoperative surgical and tumor margin assessment

“…The measured spectra can thus be analyzed accurately using diffusion theory and translated to concentrations of chromophores, such as water and fat. 28 For evaluating a larger surface, such a point measurement procedure would be unpractical and too time-consuming. Therefore, an approach based on hyperspectral imaging has been developed, focusing on the wavelength range from 950 to 1640 nm.…”