Abstract

Intraoperative fluorescence imaging informs decisions regarding surgical margins by detecting and localizing signals from fluorescent reporters, labeling targets such as malignant tissues. This guidance reduces the likelihood of undetected malignant tissue remaining after resection, eliminating the need for additional treatment or surgery. The primary challenges in performing open-air intraoperative fluorescence imaging come from the weak intensity of the fluorescence signal in the presence of strong surgical and ambient illumination, and the auto-fluorescence of non-target components, such as tissue, especially in the visible spectral window (400–650 nm). In this work, a multispectral open-air fluorescence imaging system is presented for translational image-guided intraoperative applications, which overcomes these challenges. The system is capable of imaging weak fluorescence signals with nanomolar sensitivity in the presence of surgical illumination. This is done using synchronized fluorescence excitation and image acquisition with real-time background subtraction. Additionally, the system uses a liquid crystal tunable filter for acquisition of multispectral images that are used to spectrally unmix target fluorescence from non-target auto-fluorescence. Results are validated by preclinical studies on murine models and translational canine oncology models.

© 2017 Optical Society of America

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References

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2016 (3)

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A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

N. Dimitriadis, B. Grychtol, L. Maertins, T. Behr, G. Themelis, and N. C. Deliolanis, Opt. Lett. 41, 1173 (2016).
[Crossref]

2013 (2)

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Q. T. Nguyen and R. Y. Tsien, Nat. Rev. Cancer 13, 653 (2013).
[Crossref]

2011 (1)

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

2006 (2)

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

V. Ntziachristos, Annu. Rev. Biomed. Eng. 8, 1 (2006).
[Crossref]

2005 (1)

J. M. Nascimento and J. M. Dias, IEEE Trans. Geosci. Remote Sens. 43, 898 (2005).
[Crossref]

Alencar, H.

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

Asanuma, D.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Behr, T.

Chen, X.-L.

F.-Y. Li, X.-L. Chen, and B.-N. Xu, Chronic Dis. Transl. Med. 2, 181 (2016).

Choyke, P. L.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

de Jong, R. J. B.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Deliolanis, N. C.

Dias, J. M.

J. M. Nascimento and J. M. Dias, IEEE Trans. Geosci. Remote Sens. 43, 898 (2005).
[Crossref]

Dimitriadis, N.

DSouza, A. V.

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Figueiredo, J. L.

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

Grychtol, B.

Henderson, E. R.

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Kamiya, M.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Keereweer, S.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Kerrebijn, J. D.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Kosaka, N.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Li, F.-Y.

F.-Y. Li, X.-L. Chen, and B.-N. Xu, Chronic Dis. Transl. Med. 2, 181 (2016).

Lin, H.

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Löwik, C. W.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Maertins, L.

Mahmood, U.

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

Mitsunaga, M.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Nagano, T.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Nascimento, J. M.

J. M. Nascimento and J. M. Dias, IEEE Trans. Geosci. Remote Sens. 43, 898 (2005).
[Crossref]

Nguyen, Q. T.

Q. T. Nguyen and R. Y. Tsien, Nat. Rev. Cancer 13, 653 (2013).
[Crossref]

Ntziachristos, V.

V. Ntziachristos, Annu. Rev. Biomed. Eng. 8, 1 (2006).
[Crossref]

Ogawa, M.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Pogue, B. W.

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Sakabe, M.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Samkoe, K. S.

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Snoeks, T. J.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Sterenborg, H. J.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Themelis, G.

Tsien, R. Y.

Q. T. Nguyen and R. Y. Tsien, Nat. Rev. Cancer 13, 653 (2013).
[Crossref]

Urano, Y.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Vahrmeijer, A. L.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Van Driel, P. B.

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

Weissleder, R.

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

Xu, B.-N.

F.-Y. Li, X.-L. Chen, and B.-N. Xu, Chronic Dis. Transl. Med. 2, 181 (2016).

Young, M. R.

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Annu. Rev. Biomed. Eng. (1)

V. Ntziachristos, Annu. Rev. Biomed. Eng. 8, 1 (2006).
[Crossref]

Chronic Dis. Transl. Med. (1)

F.-Y. Li, X.-L. Chen, and B.-N. Xu, Chronic Dis. Transl. Med. 2, 181 (2016).

Clin. Cancer Res. (1)

S. Keereweer, P. B. Van Driel, T. J. Snoeks, J. D. Kerrebijn, R. J. B. de Jong, A. L. Vahrmeijer, H. J. Sterenborg, and C. W. Löwik, Clin. Cancer Res. 19, 3745 (2013).

IEEE Trans. Geosci. Remote Sens. (1)

J. M. Nascimento and J. M. Dias, IEEE Trans. Geosci. Remote Sens. 43, 898 (2005).
[Crossref]

Int. J. Cancer (1)

J. L. Figueiredo, H. Alencar, R. Weissleder, and U. Mahmood, Int. J. Cancer 118, 2672 (2006).
[Crossref]

J. Biomed. Opt. (1)

A. V. DSouza, H. Lin, E. R. Henderson, K. S. Samkoe, and B. W. Pogue, J. Biomed. Opt. 21, 080901 (2016).
[Crossref]

Nat. Rev. Cancer (1)

Q. T. Nguyen and R. Y. Tsien, Nat. Rev. Cancer 13, 653 (2013).
[Crossref]

Opt. Lett. (1)

Sci. Transl. Med. (1)

Y. Urano, M. Sakabe, N. Kosaka, M. Ogawa, M. Mitsunaga, D. Asanuma, M. Kamiya, M. R. Young, T. Nagano, and P. L. Choyke, Sci. Transl. Med. 3, 110ra119 (2011).
[Crossref]

Supplementary Material (1)

NameDescription
» Visualization 1       Intraoperative fluorescence-guided tumor resection in a canine subject using Solaris.

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Figures (8)

Fig. 1.
Fig. 1.

Solaris open-air intraoperative imaging system consists of an imaging head and a base. (a) Open-air hands-free layout of Solaris that allows seamless integration into intraoperative workflows. (b) Schematic of the imaging head showing the two cameras, the lens and mirror configuration, the LED banks, and the filter wheel.

Fig. 2.
Fig. 2.

Camera acquisition and LED pulsing timing diagram illustrates the mechanism through which the stream of foreground and background frames is captured by the sCMOS camera.

Fig. 3.
Fig. 3.

Solaris open-air intraoperative fluorescence imaging. (a) Lymphatic mapping in a murine model after subcutaneous injection of AngioSense 680 EX. (b) Sensitivity studies were performed on the four channels of Solaris at video rate and f/2.5 with dilution series (1–1000 nM) of FITC, VivoTag 680, VivoTag 750, and ICG fluorescent reagents. The Michelson contrast and signal-to-noise ratio of the dilution wells were analyzed with solvent-only wells used as baseline.

Fig. 4.
Fig. 4.

Solaris multispectral imaging. (a) LCTF provides wavelength tunability using liquid crystal retarders placed between polarizers. (b) Transmission spectra of LCTF are plotted at a 10-nm pitch from 520 to 620 nm. (c) Background subtraction applied to LCTF-based multispectral images of a nu/nu mouse undergoing surgery.

Fig. 5.
Fig. 5.

Pure spectra of fluorescence sources in in vivo imaging are extracted from a multispectral image to perform spectral unmixing. (a) Three ROIs are placed on an LCTF-based multispectral image to obtain the spectra of rodent food, tissue in a murine model, and FITC. (b) Normalized spectra from the ROIs in part (a) are plotted with red, blue, and green corresponding to food, tissue, and FITC signal, respectively. (c) Spectrally unmixed components of the multispectral image in part (a) using the spectra plotted in part (b).

Fig. 6.
Fig. 6.

Tumor nodule in the peritoneum of a murine model is detected by spectral unmixing: (a) white light image, (b) fluorescence image, and (c) composite colorized unmixed image.

Fig. 7.
Fig. 7.

Spectral unmixing improves specificity and robustness of tumor detection in two FITC-based intraoperative in vivo studies on tumor-bearing murine models: (a) pre-surgery bioluminescence image of the first animal showing tumor signal with a peak radiance of 2.28 e 8    photons / s / sr / cm 2 in the upper abdominal area, (b) intraoperative fluorescence image of the first animal showing signals from the tumor area, the GI tract, skin, and abdominal tissue, (c) colorized spectrally unmixed image of the first animal, (d) pre-surgery bioluminescence image of the second animal showing signals from two tumors with peak radiances of 4e8 and 2.3 e 8    photons / s / sr / cm 2 in upper and lower abdomen, (e) intraoperative fluorescence image of the second animal with signals from tumor regions, GI tract, abdominal tissue, and skin, and (f) colorized unmixed image of the second animal.

Fig. 8.
Fig. 8.

Tumor margin identification is performed using Solaris on a canine undergoing veterinary surgery after intravenous administration of ProSense 750 FAST 24 h before surgery (see Visualization 1).

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[ I 1 I 11 ] = [ S 1 ( 1 ) S 1 ( 2 ) S 1 ( 3 ) S 11 ( 1 ) S 11 ( 2 ) S 11 ( 3 ) ] [ F 1 F 2 F 3 ] ,

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