We developed two-photon scanning patterned illumination microscopy (2P-SPIM) for super-resolution two-photon imaging. Our approach used a traditional two-photon microscopy setup with temporally modulated excitation to create patterned illumination fields. Combing nine different illuminations and structured illumination reconstruction, super-resolution imaging was achieved in two-photon microscopy. Using 2P-SPIM we achieved a lateral resolution of 141 nm, which represents an improvement by a factor of 1.9 over the corresponding diffraction limit. We further demonstrated super-resolution cellular imaging by 2P-SPIM to image actin cytoskeleton in mammalian cells and three-dimensional imaging in highly scattering retinal tissue.
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http://dx.doi.org/10.1103/PhysRevE.91.042703 | DOI Listing |
In living organisms, the natural motion caused by heartbeat, breathing, or muscle movements leads to the deformation of tissue caused by translation and stretching of the tissue structure. This effect results in the displacement or deformation of the plane of observation for intravital microscopy and causes motion-induced aberrations of the resulting image data. This, in turn, places severe limitations on the time during which specific events can be observed in intravital imaging experiments.
View Article and Find Full Text PDFBiological applications using multiphoton microscopy increasingly seek a larger field of view while maintaining sufficient temporal sampling to observe dynamic biological processes. Multiphoton imaging also requires high numerical aperture microscope objectives to realize efficient non-linear excitation and collection of fluorescence. This combination of low-magnification and high-numerical aperture poses a challenge for system design.
View Article and Find Full Text PDFAdaptive optics (AO) improves the spatial resolution of microscopy by correcting optical aberrations. While its application has been well established in microscopy modalities utilizing a circular pupil, its adaptation to systems with non-circular pupils, such as Bessel-focus two-photon fluorescence microscopy (2PFM) with an annular pupil, remains relatively uncharted. Herein, we present a modal focal AO (MFAO) method for Bessel-focus 2PFM.
View Article and Find Full Text PDFSTAR Protoc
January 2025
Department of Neurosurgery, the First Affiliated Hospital of Jinan University, Guangzhou, China; Department of Neurosurgery, the Affiliated Hospital, Southwest Medical University, Luzhou 646000, China; Laboratory of Neurological Diseases and Brain Function, the Affiliated Hospital, Southwest Medical University, Luzhou 646000, China. Electronic address:
Under pathological conditions, astrocytes can transfer mitochondria to neurons, where they exert neuroprotective effects. In this context, we present a protocol for capturing astrocytic mitochondria in neurons of adult mice using a two-photon microscope. We describe an approach for constructing a mouse model with combined labeling of astrocytic mitochondria and neurons.
View Article and Find Full Text PDFPhotosynth Res
January 2025
State Key Laboratory of Forage Breeding-by-Design and Utilization, Key Laboratory of Photobiology, Institute of Botany, Chinese Academy of Sciences, Beijing, 100093, China.
Maize (Zea mays L.) performs highly efficient C photosynthesis by dividing photosynthetic metabolism between mesophyll and bundle sheath cells. In vivo physiological measurements are indispensable for C photosynthesis research as photosynthetic activities are easily interrupted by leaf section or cell isolation.
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