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Current Protocols Jan 2023In this series of papers on light microscopy imaging, we have covered the fundamentals of microscopy, super-resolution microscopy, and lightsheet microscopy. This last... (Review)
Review
In this series of papers on light microscopy imaging, we have covered the fundamentals of microscopy, super-resolution microscopy, and lightsheet microscopy. This last review covers multi-photon microscopy with a brief reference to intravital imaging and Brainbow labeling. Multi-photon microscopy is often referred to as two-photon microscopy. Indeed, using two-photon microscopy is by far the most common way of imaging thick tissues; however, it is theoretically possible to use a higher number of photons, and three-photon microscopy is possible. Therefore, this review is titled "multi-photon microscopy." Another term for describing multi-photon microscopy is "non-linear" microscopy because fluorescence intensity at the focal spot depends upon the average squared intensity rather than the squared average intensity; hence, non-linear optics (NLO) is an alternative name for multi-photon microscopy. It is this non-linear relationship (or third exponential power in the case of three-photon excitation) that determines the axial optical sectioning capability of multi-photon imaging. In this paper, the necessity for two-photon or multi-photon imaging is explained, and the method of optical sectioning by multi-photon microscopy is described. Advice is also given on what fluorescent markers to use and other practical aspects of imaging thick tissues. The technique of Brainbow imaging is discussed. The review concludes with a description of intravital imaging of the mouse. © 2023 Wiley Periodicals LLC.
Topics: Animals; Mice; Microscopy, Fluorescence; Intravital Microscopy; Photons; Microscopy, Confocal; Optics and Photonics
PubMed: 36706245
DOI: 10.1002/cpz1.634 -
Annual Review of Biomedical Engineering 2000Two-photon fluorescence microscopy is one of the most important recent inventions in biological imaging. This technology enables noninvasive study of biological... (Review)
Review
Two-photon fluorescence microscopy is one of the most important recent inventions in biological imaging. This technology enables noninvasive study of biological specimens in three dimensions with submicrometer resolution. Two-photon excitation of fluorophores results from the simultaneous absorption of two photons. This excitation process has a number of unique advantages, such as reduced specimen photodamage and enhanced penetration depth. It also produces higher-contrast images and is a novel method to trigger localized photochemical reactions. Two-photon microscopy continues to find an increasing number of applications in biology and medicine.
Topics: Animals; Biomedical Engineering; Fluorescent Dyes; History, 20th Century; Humans; Image Processing, Computer-Assisted; Microscopy, Confocal; Microscopy, Fluorescence; Microscopy, Video; Optics and Photonics; Photons
PubMed: 11701518
DOI: 10.1146/annurev.bioeng.2.1.399 -
Lasers in Medical Science Mar 2020Deep tissue imaging using two-photon fluorescence (TPF) techniques have revolutionized the optical imaging community by providing in depth molecular information at the... (Review)
Review
Deep tissue imaging using two-photon fluorescence (TPF) techniques have revolutionized the optical imaging community by providing in depth molecular information at the single-cell level. These techniques provide structural and functional aspects of mammalian brain at unprecedented depth and resolution. However, wavefront distortions introduced by the optical system as well as the biological sample (tissue) limit the achievable fluorescence signal-to-noise ratio and resolution with penetration depth. In this review, we discuss on the advances in TPF microscopy techniques for in vivo functional imaging and offer guidelines as to which technologies are best suited for different imaging applications with special reference to adaptive optics.
Topics: Animals; Brain; Imaging, Three-Dimensional; Microscopy, Fluorescence; Neuroimaging; Optics and Photonics; Photons
PubMed: 31729608
DOI: 10.1007/s10103-019-02908-z -
Neuron Mar 1997
Review
Topics: Brain Chemistry; Calcium; Embryo, Mammalian; Embryo, Nonmammalian; Equipment Design; Forecasting; Lasers; Microscopy; Microscopy, Confocal; Microscopy, Fluorescence; Optics and Photonics; Photons
PubMed: 9115730
DOI: 10.1016/s0896-6273(00)81237-4 -
Nature Protocols Dec 2023Two-photon microscopy, combined with the appropriate optical labelling, enables the measurement and tracking of submicrometer structures within brain cells, as well as... (Review)
Review
Two-photon microscopy, combined with the appropriate optical labelling, enables the measurement and tracking of submicrometer structures within brain cells, as well as the spatiotemporal mapping of spikes in individual neurons and of neurotransmitter release in individual synapses. Yet, the spatial resolution of two-photon microscopy rapidly degrades as imaging is attempted at depths of more than a few scattering lengths into tissue, i.e., below the superficial layers that constitute the top 300-400 µm of the neocortex. To obviate this limitation, we shape the focal volume, generated by the excitation beam, by modulating the incident wavefront via guidestar-assisted adaptive optics. Here, we describe the construction, calibration and operation of a two-photon microscope that incorporates adaptive optics to restore diffraction-limited resolution at depths close to 900 µm in the mouse cortex. Our setup detects a guidestar formed by the excitation of a red-shifted dye in blood serum, used to directly measure the wavefront. We incorporate predominantly commercially available optical, optomechanical, mechanical and electronic components, and supply computer-aided design models of other customized components. The resulting adaptive optics two-photon microscope is modular and allows for expanded imaging and optical excitation capabilities. We demonstrate our methodology in the mouse neocortex by imaging the morphology of somatostatin-expressing neurons that lie 700 µm beneath the pia, calcium dynamics of layer 5b projection neurons and thalamocortical glutamate transmission to L4 neurons. The protocol requires ~30 d to complete and is suitable for users with graduate-level expertise in optics.
Topics: Mice; Animals; Microscopy; Optics and Photonics; Photons; Neurons; Calcium
PubMed: 37914781
DOI: 10.1038/s41596-023-00893-w -
Nature Nanotechnology Jun 2010The detection of photons underpins imaging, spectroscopy, fibre-optic communications and time-gated distance measurements. Nanostructured materials are attractive for... (Review)
Review
The detection of photons underpins imaging, spectroscopy, fibre-optic communications and time-gated distance measurements. Nanostructured materials are attractive for detection applications because they can be integrated with conventional silicon electronics and flexible, large-area substrates, and can be processed from the solution phase using established techniques such as spin casting, spray coating and layer-by-layer deposition. In addition, their performance has improved rapidly in recent years. Here we review progress in light sensing using nanostructured materials, focusing on solution-processed materials such as colloidal quantum dots and metal nanoparticles. These devices exhibit phenomena such as absorption of ultraviolet light, plasmonic enhancement of absorption, size-based spectral tuning, multiexciton generation, and charge carrier storage in surface and interface traps.
Topics: Nanostructures; Nanotechnology; Optics and Photonics; Photons; Surface Plasmon Resonance
PubMed: 20473301
DOI: 10.1038/nnano.2010.78 -
Optics Express Oct 2010A recently introduced two-channel confocal microscope with correlated detection promises up to 50% improvement in transverse spatial resolution [Simon, Sergienko, Optics...
A recently introduced two-channel confocal microscope with correlated detection promises up to 50% improvement in transverse spatial resolution [Simon, Sergienko, Optics Express 18, 9765 (2010)] via the use of photon correlations. Here we achieve similar results in a different manner, introducing a triple-confocal correlated microscope which exploits the correlations present in optical parametric amplifiers. It is based on tight focusing of pump radiation onto a thin sample positioned in front of a nonlinear crystal, followed by coincidence detection of signal and idler photons, each focused onto a pinhole. This approach offers further resolution enhancement in confocal microscopy.
Topics: Algorithms; Amplifiers, Electronic; Equipment Design; Microscopy, Confocal; Models, Statistical; Models, Theoretical; Optics and Photonics; Oscillometry; Photons
PubMed: 20941116
DOI: 10.1364/OE.18.022147 -
Nature Communications Jun 2022When pursuing sustainable approaches to fabricate photonic structures, nature can be used as a source of inspiration for both the nanoarchitecture and the constituent...
When pursuing sustainable approaches to fabricate photonic structures, nature can be used as a source of inspiration for both the nanoarchitecture and the constituent materials. Although several biomaterials have been promised as suitable candidates for photonic materials and pigments, their fabrication processes have been limited to the small to medium-scale production of films. Here, by employing a substrate-free process, structurally coloured microparticles are produced via the confined self-assembly of a cholesteric cellulose nanocrystal (CNC) suspension within emulsified microdroplets. Upon drying, the droplets undergo multiple buckling events, which allow for greater contraction of the nanostructure than predicted for a spherical geometry. This buckling, combined with a solvent or thermal post-treatment, enables the production of dispersions of vibrant red, green, and blue cellulose photonic pigments. The hierarchical structure of these pigments enables the deposition of coatings with angular independent colour, offering a consistent visual appearance across a wide range of viewing angles.
Topics: Cellulose; Nanoparticles; Nanostructures; Optics and Photonics; Photons
PubMed: 35697688
DOI: 10.1038/s41467-022-31079-9 -
Biotechnology Progress 1997The ability to map interior optical properties of a highly scattering medium from exterior measurements of light propagation is afforded by optical tomography. In this... (Review)
Review
The ability to map interior optical properties of a highly scattering medium from exterior measurements of light propagation is afforded by optical tomography. In this communication, we describe the problem of optical tomography, the techniques of photon migration measurements necessary to accomplish it, and the development of multipixel measurements for rapid collection of optical signals. These multipixel measurements are shown to provide detection of contrast owing to the optical properties of absorption and fluorescence associated with dye-laden heterogeneities embedded in a tissue-like scattering medium. From these rapid measurements, successful reconstruction of an interior optical property map may now be possible with clinically realistic data acquisition times. Applications for the technology arise for biomedical optical imaging for the in vivo detection of disease and the diagnosis of tissue (bio-) chemistry.
Topics: Absorption; Fluorescence; Mathematics; Optics and Photonics; Photons; Tomography
PubMed: 9336987
DOI: 10.1021/bp970085g -
Nature Communications Dec 2019Constructing colloidal particles into functional nanostructures, materials, and devices is a promising yet challenging direction. Many optical techniques have been...
Constructing colloidal particles into functional nanostructures, materials, and devices is a promising yet challenging direction. Many optical techniques have been developed to trap, manipulate, assemble, and print colloidal particles from aqueous solutions into desired configurations on solid substrates. However, these techniques operated in liquid environments generally suffer from pattern collapses, Brownian motion, and challenges that come with reconfigurable assembly. Here, we develop an all-optical technique, termed optothermally-gated photon nudging (OPN), for the versatile manipulation and dynamic patterning of a variety of colloidal particles on a solid substrate at nanoscale accuracy. OPN takes advantage of a thin surfactant layer to optothermally modulate the particle-substrate interaction, which enables the manipulation of colloidal particles on solid substrates with optical scattering force. Along with in situ optical spectroscopy, our non-invasive and contactless nanomanipulation technique will find various applications in nanofabrication, nanophotonics, nanoelectronics, and colloidal sciences.
Topics: Biophysical Phenomena; Colloids; Motion; Nanostructures; Optics and Photonics; Particle Size; Photons; Surface-Active Agents; Temperature
PubMed: 31831746
DOI: 10.1038/s41467-019-13676-3