Label-free concurrent 5-modal microscopy (Co5M) resolves unknown spatio-temporal processes in wound healing

• Published in: Communications biology 2021-09, Vol.4 (1), p.1040-1040, Article 1040
• Main Authors: Seeger, Markus, Dehner, Christoph, Jüstel, Dominik, Ntziachristos, Vasilis
• Format: Article
• Language: English
• Subjects: 14/10; 14/63; 14/69; 59; 631/1647/245; 631/1647/328; 631/57; 64/60; 692/53; Angiogenesis; Animals; Biology; Biomedical and Life Sciences; Collagen; Female; Life Sciences; Mice; Mice, Nude; Microscopy; Microscopy - instrumentation; Microvasculature; Plasticity; Sebaceous gland; Spatio-Temporal Analysis; Vasodilation; Wound Healing
• ISSN: 2399-3642; 2399-3642
• DOI: 10.1038/s42003-021-02573-5

The non-invasive investigation of multiple biological processes remains a methodological challenge as it requires capturing different contrast mechanisms, usually not available with any single modality. Intravital microscopy has played a key role in dynamically studying biological morphology and function, but it is generally limited to resolving a small number of contrasts, typically generated by the use of transgenic labels, disturbing the biological system. We introduce concurrent 5-modal microscopy (Co5M), illustrating a new concept for label-free in vivo observations by simultaneously capturing optoacoustic, two-photon excitation fluorescence, second and third harmonic generation, and brightfield contrast. We apply Co5M to non-invasively visualize multiple wound healing biomarkers and quantitatively monitor a number of processes and features, including longitudinal changes in wound shape, microvascular and collagen density, vessel size and fractality, and the plasticity of sebaceous glands. Analysis of these parameters offers unique insights into the interplay of wound closure, vasodilation, angiogenesis, skin contracture, and epithelial reformation in space and time, inaccessible by other methods. Co5M challenges the conventional concept of biological observation by yielding multiple simultaneous parameters of pathophysiological processes in a label-free mode. Seeger et al. demonstrate concurrent 5-modal microscopy (Co5M), combining optoacoustic, two-photon, second and third harmonic generation, and brightfield microscopy in one system. Longitudinal changes associated with the wound healing process in mouse ears were monitored label-free in vivo using complementary information obtained from the Co5M system to characterize wound shape, microvascular and collagen density, vessel size and fractality, and the plasticity of sebaceous glands.