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Biomed Opt Express. 2018 Apr 30;9(5):2457-2475. doi: 10.1364/BOE.9.002457. eCollection 2018 May 1.

Multiscale nonlinear microscopy and widefield white light imaging enables rapid histological imaging of surgical specimen margins.

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Massachusetts Institute of Technology, Department of Electrical Engineering and Computer Science and Research Laboratory of Electronics, 32 Vassar Street, Cambridge, MA 02139,USA.
Harvard Medical School, Department of Pathology, Beth Israel Deaconess Medical Center, 330 Brookline Avenue, Boston, MA 02215, USA.
Thorlabs Advanced Imaging Group, 108 Powers Court, Sterling, VA 20166, USA.


The ability to histologically assess surgical specimens in real-time is a long-standing challenge in cancer surgery, including applications such as breast conserving therapy (BCT). Up to 40% of women treated with BCT for breast cancer require a repeat surgery due to postoperative histological findings of close or positive surgical margins using conventional formalin fixed paraffin embedded histology. Imaging technologies such as nonlinear microscopy (NLM), combined with exogenous fluorophores can rapidly provide virtual H&E imaging of surgical specimens without requiring microtome sectioning, facilitating intraoperative assessment of margin status. However, the large volume of typical surgical excisions combined with the need for rapid assessment, make comprehensive cellular resolution margin assessment during surgery challenging. To address this limitation, we developed a multiscale, real-time microscope with variable magnification NLM and real-time, co-registered position display using a widefield white light imaging system. Margin assessment can be performed rapidly under operator guidance to image specific regions of interest located using widefield imaging. Using simulated surgical margins dissected from human breast excisions, we demonstrate that multi-centimeter margins can be comprehensively imaged at cellular resolution, enabling intraoperative margin assessment. These methods are consistent with pathology assessment performed using frozen section analysis (FSA), however NLM enables faster and more comprehensive assessment of surgical specimens because imaging can be performed without freezing and cryo-sectioning. Therefore, NLM methods have the potential to be applied to a wide range of intra-operative applications.


(170.2520) Fluorescence microscopy; (170.4580) Optical diagnostics for medicine; (170.4730) Optical pathology; (180.4315) Nonlinear microscopy

Conflict of interest statement

The authors have filed intellectual property on portions of the methods and apparatus described in this article.

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