Dual Near‐Infrared Two‐Photon Microscopy for Deep‐Tissue Dopamine Nanosensor Imaging |
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Authors: | Jackson T Del Bonis‐O'Donnell Ralph H Page Abraham G Beyene Eric G Tindall Ian R McFarlane Markita P Landry |
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Affiliation: | 1. Chemical and Biomolecular Engineering, University of California, Berkeley, CA, USA;2. California Institute for Quantitative Biosciences, QB3, University of California, Berkeley, CA, USA;3. Chan‐Zuckerberg Biohub, San Francisco, CA, USA |
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Abstract: | A key limitation for achieving deep imaging in biological structures lies in photon absorption and scattering leading to attenuation of fluorescence. In particular, neurotransmitter imaging is challenging in the biologically relevant context of the intact brain for which photons must traverse the cranium, skin, and bone. Thus, fluorescence imaging is limited to the surface cortical layers of the brain, only achievable with craniotomy. Herein, this study describes optimal excitation and emission wavelengths for through‐cranium imaging, and demonstrates that near‐infrared emissive nanosensors can be photoexcited using a two‐photon 1560 nm excitation source. Dopamine‐sensitive nanosensors can undergo two‐photon excitation, and provide chirality‐dependent responses selective for dopamine with fluorescent turn‐on responses varying between 20% and 350%. The two‐photon absorption cross‐section and quantum yield of dopamine nanosensors are further calculated, and a two‐photon power law relationship for the nanosensor excitation process is confirmed. Finally, the improved image quality of the nanosensors embedded 2‐mm‐deep into a brain‐mimetic tissue phantom is shown, whereby one‐photon excitation yields 42% scattering, in contrast to 4% scattering when the same object is imaged under two‐photon excitation. The approach overcomes traditional limitations in deep‐tissue fluorescence microscopy, and can enable neurotransmitter imaging in the biologically relevant milieu of the intact and living brain. |
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Keywords: | deep‐tissue microscopy dopamine nanosensors near‐infrared fluorescent sensors single‐walled carbon nanotubes two‐photon microscopy |
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