Fluorescence imaging has revolutionized biomedical research over the past three decades. Its high molecular specificity and unrivalled single-molecule-level sensitivity have enabled breakthroughs in a number of research fields. For in vivo applications its major limitation is its superficial imaging depth, a result of random scattering in biological tissues causing exponential attenuation of the ballistic component of a light wave. Here, we present fluorescence imaging beyond the ballistic regime by combining single-cycle pulsed ultrasound modulation and digital optical phase conjugation. We demonstrate a near-isotropic three-dimensional localized sound–light interaction zone. With the exceptionally high optical gain provided by the digital optical phase conjugation system, we can deliver sufficient optical power to a focus inside highly scattering media for not only fluorescence imaging but also a variety of linear and nonlinear spectroscopy measurements. This technology paves the way for many important applications in both fundamental biology research and clinical studies.
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The authors thank C. Shank, Y.M. Wang and C. Yang for helpful discussions, T.-W. Chen for instructions on the micropipette puller and A. Hu for preparing the fixed rat brain slices. The research was supported by the Howard Hughes Medical Institute.
The authors declare no competing financial interests.
About this article
Journal of Biophotonics (2019)
Journal of Physics D: Applied Physics (2019)
Scientific Reports (2019)
Optics Express (2019)