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A 3-chip CCD imaging system has been developed for quantitative in vivo fluorescence imaging. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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However, due to the infiltrative nature of most of the gliomas, complete resection is difficult to achieve, resulting in high risk of tumor recurrence [2].

Hence, more sensitive and specific techniques are needed to aid in the identification of malignant tissue intraoperatively and, thereby, to increase the completeness of tumor resection without damaging adjacent critical normal brain structures and function [4].

Spectral response curves for the red, green, and blue channels of the CCD were measured using a standard color target (Color Checker Chart: Gretag Macbeth, Grand Rapids, MI, USA) illuminated by a tungsten-halogen light source (LS-1: Ocean Optics, Dunedin, FL, USA).

For this, the images of the color target were taken through the laparoscope without the fluorescence emission filter installed.

This has been validated in phantoms for a wide range of fluorophore concentrations (in the case of Pp IX, e.g., to ) and tissue optical properties [41].

Here, we present a prototype FGR instrument that provides video-rate digital fluorescence imaging incorporating a double-ratiometric correction method that is optimized for intraoperative identification of brain tumors.

In the simplest approach, correction is performed using single excitation and emission wavelengths [34, 35], but this does not correct for tissue autofluorescence [36].

As we recently showed in a modeling study [37], the use of multiple excitation and/or emission wavelengths can virtually eliminate the tissue autofluorescence and also enable quantitative measurements of fluorophores [36–40], such that the fluorescent signal depends only on the concentration of the fluorophore.

For the present study, the excitation filters had central wavelengths, , at the Pp IX Soret maxima of 405 nm (full width at half maximum, FWHM = 96 nm) and at 440 nm (FWHM = 60 nm).

The latter was chosen as the shortest wavelength lying above the main Pp IX Soret band in order to allow for correction for the tissue autofluorescence.

A long-pass 500 nm filter (Custom made, Chroma Technologies, VT, USA) is placed between the camera and the laparoscope.

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