Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Two-photon imaging of the immune system.

ABSTRACT: Two-photon microscopy is a powerful method for visualizing biological processes as they occur in their native environment in real time. The immune system uniquely benefits from this technology as most of its constituent cells are highly motile and interact extensively with each other and with the environment. Two-photon microscopy has provided many novel insights into the dynamics of the development and function of the immune system that could not have been deduced by other methods and has become an indispensible tool in the arsenal of immunologists. In this unit, we provide several protocols for preparation of various organs for imaging by two-photon microscopy that are intended to introduce the new user to some basic aspects of this method.

SUBMITTER: Dzhagalov IL

PROVIDER: S-EPMC3662370 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Two-photon imaging of the immune system.

Dzhagalov Ivan L IL Melichar Heather J HJ Ross Jenny O JO Herzmark Paul P Robey Ellen A EA

Current protocols in cytometry 20120401

Two-photon microscopy is a powerful method for visualizing biological processes as they occur in their native environment in real time. The immune system uniquely benefits from this technology as most of its constituent cells are highly motile and interact extensively with each other and with the environment. Two-photon microscopy has provided many novel insights into the dynamics of the development and function of the immune system that could not have been deduced by other methods and has becom ...[more]

PMID: 22470153

Similar Datasets

tRNA analysis and two-photon imaging of a 5,000-year-old human femur

Project description:We report the application of tRNA profiling in an ancient sample, demonstrating the use of RNA in archeogenomics.

2021-08-13 | GSE85722 | GEO

Video-rate two-photon excited fluorescence lifetime imaging system with interleaved digitization.

Project description:A fast (up to video rate) two-photon excited fluorescence lifetime imaging system based on interleaved digitization is demonstrated. The system is compatible with existing beam-scanning microscopes with minor electronics and software modification. Proof-of-concept demonstrations were performed using laser dyes and biological tissue.

| S-EPMC4756458 | biostudies-literature

A silent two-photon imaging system for studying in vivo auditory neuronal functions.

Project description:Two-photon laser-scanning microscopy has become an essential tool for imaging neuronal functions in vivo and has been applied to different parts of the neural system, including the auditory system. However, many components of a two-photon microscope, such as galvanometer-based laser scanners, generate mechanical vibrations and thus acoustic artifacts, making it difficult to interpret auditory responses from recorded neurons. Here, we report the development of a silent two-photon imaging system and its applications in the common marmoset (Callithrix Jacchus), a non-human primate species sharing a similar hearing range with humans. By utilizing an orthogonal pair of acousto-optical deflectors (AODs), full-frame raster scanning at video rate was achieved without introducing mechanical vibrations. Imaging depth can be optically controlled by adjusting the chirping speed on the AODs without any mechanical motion along the Z-axis. Furthermore, all other sound-generating components of the system were acoustically isolated, leaving the noise floor of the working system below the marmoset's hearing threshold. Imaging with the system in awake marmosets revealed many auditory cortex neurons that exhibited maximal responses at low sound levels, which were not possible to study using traditional two-photon imaging systems. This is the first demonstration of a silent two-photon imaging system that is capable of imaging auditory neuronal functions in vivo without acoustic artifacts. This capacity opens new opportunities for a better understanding of auditory functions in the brain and helps isolate animal behavior from microscope-generated acoustic interference.

| S-EPMC9010453 | biostudies-literature

Denoising two-photon calcium imaging data.

Project description:Two-photon calcium imaging is now an important tool for in vivo imaging of biological systems. By enabling neuronal population imaging with subcellular resolution, this modality offers an approach for gaining a fundamental understanding of brain anatomy and physiology. Proper analysis of calcium imaging data requires denoising, that is separating the signal from complex physiological noise. To analyze two-photon brain imaging data, we present a signal plus colored noise model in which the signal is represented as harmonic regression and the correlated noise is represented as an order autoregressive process. We provide an efficient cyclic descent algorithm to compute approximate maximum likelihood parameter estimates by combing a weighted least-squares procedure with the Burg algorithm. We use Akaike information criterion to guide selection of the harmonic regression and the autoregressive model orders. Our flexible yet parsimonious modeling approach reliably separates stimulus-evoked fluorescence response from background activity and noise, assesses goodness of fit, and estimates confidence intervals and signal-to-noise ratio. This refined separation leads to appreciably enhanced image contrast for individual cells including clear delineation of subcellular details and network activity. The application of our approach to in vivo imaging data recorded in the ferret primary visual cortex demonstrates that our method yields substantially denoised signal estimates. We also provide a general Volterra series framework for deriving this and other signal plus correlated noise models for imaging. This approach to analyzing two-photon calcium imaging data may be readily adapted to other computational biology problems which apply correlated noise models.

| S-EPMC3110192 | biostudies-literature

Two-photon imaging of soliton dynamics.

Project description:Optical solitary waves (solitons) that interact in a nonlinear system can bind and form a structure similar to a molecule. The rich dynamics of this process have created a demand for rapid spectral characterization to deepen the understanding of soliton physics with many practical implications. Here, we demonstrate stroboscopic, two-photon imaging of soliton molecules (SM) with completely unsynchronized lasers, where the wavelength and bandwidth constraints are considerably eased compared to conventional imaging techniques. Two-photon detection enables the probe and tested oscillator to operate at completely different wavelengths, which permits mature near-infrared laser technology to be leveraged for rapid SM studies of emerging long-wavelength laser sources. As a demonstration, using a 1550 nm probe laser we image the behavior of soliton singlets across the 1800-2100 nm range, and capture the rich dynamics of evolving multiatomic SM. This technique may prove to be an essential, easy-to-implement diagnostic tool for detecting the presence of loosely-bound SM, which often remain unnoticed due to instrumental resolution or bandwidth limitations.

| S-EPMC10247745 | biostudies-literature

Simultaneous two-photon imaging and two-photon optogenetics of cortical circuits in three dimensions.

Project description:The simultaneous imaging and manipulating of neural activity could enable the functional dissection of neural circuits. Here we have combined two-photon optogenetics with simultaneous volumetric two-photon calcium imaging to measure and manipulate neural activity in mouse neocortex in vivo in three-dimensions (3D) with cellular resolution. Using a hybrid holographic approach, we simultaneously photostimulate more than 80 neurons over 150 μm in depth in layer 2/3 of the mouse visual cortex, while simultaneously imaging the activity of the surrounding neurons. We validate the usefulness of the method by photoactivating in 3D selected groups of interneurons, suppressing the response of nearby pyramidal neurons to visual stimuli in awake animals. Our all-optical approach could be used as a general platform to read and write neuronal activity.

| S-EPMC5832414 | biostudies-literature

Longitudinal in vivo two-photon fluorescence imaging.

Project description:Fluorescence microscopy is an essential technique for the basic sciences, especially biomedical research. Since the invention of laser scanning confocal microscopy in the 1980s, which enabled imaging both fixed and living biological tissue with 3D precision, high-resolution fluorescence imaging has revolutionized biological research. Confocal microscopy, by its very nature, has one fundamental limitation. Due to the confocal pinhole, deep tissue fluorescence imaging is not practical. In contrast (no pun intended), two-photon fluorescence microscopy allows, in principle, the collection of all emitted photons from fluorophores in the imaged voxel, dramatically extending our ability to see deep into living tissue. Since the development of transgenic mice with genetically encoded fluorescent protein in neocortical cells in 2000, two-photon imaging has enabled the dynamics of individual synapses to be followed for up to 2 years. Since the initial landmark contributions to this field in 2002, the technique has been used to understand how neuronal structure are changed by experience, learning, and memory and various diseases. Here we provide a basic summary of the crucial elements that are required for such studies, and discuss many applications of longitudinal two-photon fluorescence microscopy that have appeared since 2002.

| S-EPMC4136386 | biostudies-literature

Two-photon imaging of the trabecular meshwork.

Project description:PurposeTo image the trabecular meshwork (TM) in its native unfixed state using a non-invasive, non-destructive technique.MethodsTwo-photon microscopy (2PM), including two-photon excitation fluorescence (2PEF) and second harmonic generation (SHG), was used to image flat-mounted trabecular meshwork samples from human cadaver eyes. Multiple images were analyzed along the tissue axis (z-axis) to generate a three-dimensional (3D) model of the region.ResultsA lattice of large collagen fibers (approximately 10 microm in diameter) were detected by inherent fluorescence (2PEF) and SHG. There are regions of both tightly overlapping bundles as well as fluid-filled regions visible from the surface of the TM. 3D analysis of multiple images reveals that the open regions deep in the TM penetrate the juxtacanalicular TM (JTM) and connect to the inner wall of Schlemm's canal (IWSC). These open regions may represent low-resistance fluid pathways between the anterior chamber and Schlemm's canal (SC).Conclusions2PM imaging of the outflow system of the human eye documented collagenous structures solely from inherent optical properties, without addition of an exogenous fluorescent label. 2PM successfully imaged into the TM without the need for fixation, embedding, or histological processing. Deep penetration using advanced optical techniques revealed regions likely representing pores in the IWSC that have been documented by multiple electron microscope studies. Our work reveals that 2PM imaging has potential as a new metric for evaluating the aqueous outflow region of the human eye and is worthy of further exploration.

| S-EPMC2890557 | biostudies-literature

High contrast two-photon imaging of fingermarks.

Project description:Optically-acquired fingermarks are widely used as evidence across law enforcement agencies as well as in the courts of law. A common technique for visualizing latent fingermarks on nonporous surfaces consists of cyanoacrylate fuming of the fingerprint material, followed by impregnation with a fluorescent dye, which under ultra violet (UV) illumination makes the fingermarks visible and thus accessible for digital recording. However, there exist critical circumstances, when the image quality is compromised due to high background scattering, high auto-fluorescence of the substrate material, or other detrimental photo-physical and photo-chemical effects such as light-induced damage to the sample. Here we present a novel near-infrared (NIR), two-photon induced fluorescence imaging modality, which significantly enhances the quality of the fingermark images, especially when obtained from highly reflective and/or scattering surfaces, while at the same time reducing photo-damage to sensitive forensic samples.

| S-EPMC4823789 | biostudies-literature

Optical constraints on two-photon voltage imaging.

Project description:Genetically encoded voltage indicators (GEVIs) are a valuable tool for studying neural circuits in vivo, but the relative merits of one-photon (1P) vs. two-photon (2P) voltage imaging are not well characterized. Here we compare the photophysical and imaging properties of commonly used GEVIs under 1P and 2P excitation. 2P excitation requires ~104-fold more illumination power per cell to produce comparable photon count rates to 1P excitation, driving a stringent tradeoff between shot noise and tissue photodamage.

| S-EPMC10680948 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data