A paradigm shift for 3D optical imaging of biological tissues

Non-invasive microscopy exploits light scattering to obtain a three-dimensional image of biological tissues. However, light propagation is drastically affected by the heterogeneities of tissues, which totally blurs microscopic images in depth. Adapted from astronomy, adaptive optics has been developed to compensate for these aberrations in microscopy, but the associated frame rate and penetration depth remain extremely limited for 3D imaging. We have therefore developed a revolutionary microscope that can acquire three-dimensional images of tissue at millimetric depths, with sub-cellular resolution (250 nm) and high frame rates (1 to 200 Hz).
This paradigm shift is based on ultra-fast acquisition of the reflection matrix using sparse sample illumination and interferometric measurement of reflected light at multiple wavelengths. The light focusing process is optimized in post-processing for any voxel in the medium, using smart algorithms to overcome the problems of aberrations and multiple scattering that have hitherto impaired the performance of conventional microscopes. Matrix imaging is also the subject of an article published in Nature Communications, demonstrating the significant extension of penetration depth in optical coherence tomography thanks to the harnessing of multiple scattering paths.

 Figure: a Multichromatic and multiplexed illumination of the sample allowing ultra-fast acquisition of the multi-spectral reflection matrix. b Standard 3D image of the cornea blurred by aberrations and multiple scattering. c Matrix image of the cornea revealing the 3D arrangement of structures invisible on the initial image, such as nerves and corneal fibroblasts sandwiched between layers of collagen lamellae. Image credit: Victor Barolle and Flavien Bureau
These works pave the way for a fully digital microscope enabling quantitative, in-depth in-vivo tissue inspection in real time. Applications range from the control of embryos in the in-vitro fertilization process to the control of organoids for biomedical applications, ophthalmology (high-resolution imaging of the cornea and ultra-deep imaging of the retina) and dermatology (in-depth control of melanomas). 
Paul Balondrade, Victor Barolle, Nicolas Guigui & Alexandre Aubry