Multiphoton Multispectral Fluorescence Lifetime Tomography for the Evaluation of Basal Cell Carcinomas

Abstract

We present the first detailed study using multispectral multiphoton fluorescence lifetime imaging to differentiate basal cell carcinoma cells (BCCs) from normal keratinocytes. Images were acquired from 19 freshly excised BCCs and 27 samples of normal skin (in & ex vivo). Features from fluorescence lifetime images were used to discriminate BCCs with a sensitivity/specificity of 79%/93% respectively. A mosaic of BCC fluorescence lifetime images covering >1 mm2 is also presented, demonstrating the potential for tumour margin delineation. Using 10,462 manually segmented cells from the image data, we quantify the cellular morphology and spectroscopic differences between BCCs and normal skin for the first time. Statistically significant increases were found in the fluorescence lifetimes of cells from BCCs in all spectral channels, ranging from 19.9% (425–515 nm spectral emission) to 39.8% (620–655 nm emission). A discriminant analysis based diagnostic algorithm allowed the fraction of cells classified as malignant to be calculated for each patient. This yielded a receiver operator characteristic area under the curve for the detection of BCC of 0.83. We have used both morphological and spectroscopic parameters to discriminate BCC from normal skin, and provide a comprehensive base for how this technique could be used for BCC assessment in clinical practice.

A flexible wide‐field FLIM endoscope utilising blue excitation light for label‐free contrast of tissue

“…[12], and in humans, e.g. [13] where studies of skin autofluorescence parameters including lifetime have been correlated with cancer [14,15]. We are interested to apply FLIM to detect and monitor disease in internal organs such as the colon, for which flexible endoscope-based-instrumentation is required and where we believe that FLIM could complement current techniques such as white light endoscopy.…”

Fluorescence lifetime imaging (FLIM): Basic concepts and some recent developments

“…FLIM of autofluorescence has potential as a label-free clinical diagnostic tool for in vivo optical biopsies, in particular for skin (Dancik et al, 2013;Galletly et al, 2008;Patalay et al, 2012;Sanchez et al, 2010Sanchez et al, , 2012Seidenari et al, 2012). These 3-dimensional optical biopsies do not require any removal of tissue samples, or any other mechanical or chemical treatment.…”

Novel Approaches to Imaging Basal Cell Carcinoma

“…Fluorescence intensity and FLIM images were collected from 27 normal skin samples and 19 excised BCCs, using four emission spectral channels: blue, green, yellow and red. They reported that with this technique, detecting BCCs was accomplished with a sensitivity and specificity of 79 and 93%, respectively [19].…”