Article

Stain-free histopathology by programmable supercontinuum pulses

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Abstract

The preparation, staining, visualization and interpretation of histological images of tissue is well accepted as the gold standard process for the diagnosis of disease. These methods have a long history of development, and are used ubiquitously in pathology, despite being highly time- and labour-intensive. Here, we introduce a unique optical imaging platform and methodology for label-free multimodal multiphoton microscopy that uses a novel photonic-crystal fibre source to generate tailored chemical contrast based on programmable supercontinuum pulses. We demonstrate the collection of optical signatures of the tumour microenvironment, including evidence of mesoscopic biological organization, tumour cell migration and (lymph-) angiogenesis collected directly from fresh ex vivo mammary tissue. Acquisition of these optical signatures and other cellular or extracellular features, which are largely absent from histologically processed and stained tissue, combined with an adaptable platform for optical alignment-free programmable-contrast imaging, offers the potential to translate stain-free molecular histopathology into routine clinical use.

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Acknowledgements

This research was supported by grants from the National Institutes of Health (R01 CA166309 and R01 EB013723), the Danish Council for Independent Research – Technology and Production Sciences (FTP project ALFIE), the European Commission (EU Career Integration Grant 334324 LIGHTER) and by the Max Planck Society.

Author information

Author notes

    • Haohua Tu
    •  & Yuan Liu

    These authors contributed equally to this work

Affiliations

  1. Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA

    • Haohua Tu
    • , Yuan Liu
    • , Marina Marjanovic
    • , Eric J. Chaney
    • , Youbo Zhao
    • , Sixian You
    •  & Stephen A. Boppart
  2. Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA

    • Yuan Liu
    • , Marina Marjanovic
    • , Sixian You
    •  & Stephen A. Boppart
  3. Max Planck Institute for Polymer Research, Ackermannweg 10, Mainz 55128, Germany

    • Dmitry Turchinovich
  4. NKT Photonics A/S, Blokken 84, 3460 Birkerød, Denmark

    • Jens K. Lyngsø
  5. DTU Fotonik, Technical University of Denmark, Ørsteds Plads 343, 2800 Lyngby, Denmark

    • Jesper Lægsgaard
  6. Department of Materials Science and Engineering, Frederick Seitz Materials Research Laboratory, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA

    • William L. Wilson
  7. Biophotonic Solutions Inc., East Lansing, Michigan 48823, USA

    • Bingwei Xu
  8. Department of Chemistry and Department of Physics, Michigan State University, East Lansing, Michigan 48824, USA

    • Marcos Dantus
  9. Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA

    • Stephen A. Boppart
  10. College of Medicine, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA

    • Stephen A. Boppart

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Contributions

H.T., Y.L. and S.A.B. conceived the idea, performed the analysis and wrote the manuscript. H.T., Y.L. and Y.Z. built the microscope and conducted optical experiments. H.T., D.T. and J.L. tested and improved the long-term stability of the fibre supercontinuum source. J.K.L. fabricated a variety of photonic-crystal fibres for the supercontinuum source. W.L.W. characterized the supercontinuum source. E.J.C., S.Y. and M.M. performed biological experiments. B.X. and M.D. built the pulse shaper. H.T. and S.A.B. obtained funding for this research.

Competing interests

M.D. is the founder of Biophotonic Solutions Inc., and has financial interest in a proprietary pulse shaping technology using multiphoton intrapulse interference phase scan (MIIPS).

Corresponding authors

Correspondence to Haohua Tu or Stephen A. Boppart.

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