Browse
Submit Data
Databases
API
Help

Dataset Information

34 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

3D second harmonic generation imaging tomography by multi-view excitation.

ABSTRACT: Biological tissues have complex 3D collagen fiber architecture that cannot be fully visualized by conventional second harmonic generation (SHG) microscopy due to electric dipole considerations. We have developed a multi-view SHG imaging platform that successfully visualizes all orientations of collagen fibers. This is achieved by rotating tissues relative to the excitation laser plane of incidence, where the complete fibrillar structure is then visualized following registration and reconstruction. We evaluated high frequency and Gaussian weighted fusion reconstruction algorithms, and found the former approach performs better in terms of the resulting resolution. The new approach is a first step toward SHG tomography.

SUBMITTER: Campbell KR

PROVIDER: S-EPMC5847324 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

3D second harmonic generation imaging tomography by multi-view excitation.

Campbell Kirby R KR Wen Bruce B Shelton Emily M EM Swader Robert R Cox Benjamin L BL Eliceiri Kevin K Campagnola Paul J PJ

Optica 20170928 10

Biological tissues have complex 3D collagen fiber architecture that cannot be fully visualized by conventional second harmonic generation (SHG) microscopy due to electric dipole considerations. We have developed a multi-view SHG imaging platform that successfully visualizes all orientations of collagen fibers. This is achieved by rotating tissues relative to the excitation laser plane of incidence, where the complete fibrillar structure is then visualized following registration and reconstructio ...[more]

PMID: 29541654

Similar Datasets

Efficient second-harmonic imaging of collagen in histological slides using Bessel beam excitation.

Project description:Second-harmonic generation (SHG) is the most specific label-free indicator of collagen accumulation in widespread pathologies such as fibrosis, and SHG-based measurements hold important potential for biomedical analyses. However, efficient collagen SHG scoring in histological slides is hampered by the limited depth-of-field of usual nonlinear microscopes relying on focused Gaussian beam excitation. In this work we analyze theoretically and experimentally the use of Bessel beam excitation to address this issue. Focused Bessel beams can provide an axially extended excitation volume for nonlinear microscopy while preserving lateral resolution. We show that shaping the focal volume has consequences on signal level and scattering directionality in the case of coherent signals (such as SHG) which significantly differ from the case of incoherent signals (two-photon excited fluorescence, 2PEF). We demonstrate extended-depth SHG-2PEF imaging of fibrotic mouse kidney histological slides. Finally, we show that Bessel beam excitation combined with spatial filtering of the harmonic light in wave vector space can be used to probe collagen accumulation more efficiently than the usual Gaussian excitation scheme. These results open the way to SHG-based histological diagnoses.

| S-EPMC4951755 | biostudies-literature

High-Resolution Plasma Membrane-Selective Imaging by Second Harmonic Generation.

Project description:The plasma membrane is the site of intercellular communication and subsequent intracellular signal transduction. The specific visualization of the plasma membrane in living cells, however, is difficult using fluorescence-based techniques owing to the high background signals from intracellular organelles. In this study, we show that second harmonic generation (SHG) is a high-resolution plasma membrane-selective imaging technique that enables multifaceted investigations of the plasma membrane. In contrast to fluorescence imaging, SHG specifically visualizes the plasma membrane at locations that are not attached to artificial substrates and allows high-resolution imaging because of its subresolution nature. These properties were exploited to measure the distances from the plasma membrane to subcortical actin and tubulin fibers, revealing the precise cytoskeletal organization beneath the plasma membrane. Thus, SHG imaging enables the specific visualization of phenomena at the plasma membrane with unprecedented precision and versatility and should facilitate cell biology research focused on the plasma membrane.

| S-EPMC6249386 | biostudies-literature

Second Harmonic Generation Imaging Analysis of Collagen Arrangement in Human Cornea.

Project description:PurposeTo describe the horizontal arrangement of human corneal collagen bundles by using second harmonic generation (SHG) imaging.MethodsHuman corneas were imaged with an inverted two photon excitation fluorescence microscope. The excitation laser (Ti:Sapphire) was tuned to 850 nm. Backscatter signals of SHG were collected through a 425/30-nm bandpass emission filter. Multiple, consecutive, and overlapping image stacks (z-stacks) were acquired to generate three dimensional data sets. ImageJ software was used to analyze the arrangement pattern (irregularity) of collagen bundles at each image plane.ResultsCollagen bundles in the corneal lamellae demonstrated a complex layout merging and splitting within a single lamellar plane. The patterns were significantly different in the superficial and limbal cornea when compared with deep and central regions. Collagen bundles were smaller in the superficial layer and larger in deep lamellae.ConclusionsBy using SHG imaging, the horizontal arrangement of corneal collagen bundles was elucidated at different depths and focal regions of the human cornea.

| S-EPMC4553931 | biostudies-literature

Deciphering second harmonic generation signals.

Project description:Second harmonic generation (SHG) has emerged as one of the most powerful techniques used to selectively monitor surface dynamics and reactions for all types of interfaces as well as for imaging non-centrosymmetric structures, although the molecular origin of the SHG signal is still poorly understood. Here, we present a breakthrough approach to predict and interpret the SHG signal at the atomic level, which is freed from the hyperpolarisability concept and self-consistently considers the non-locality and the coupling with the environment. The direct ab initio method developed here shows that a bulk quadrupole contribution significantly overwhelms the interface dipole term in the purely interfacial induced second-order polarisation for water/air interfaces. The obtained simulated SHG responses are in unprecedented agreement with the experimental signal. This work not only paves the road for the prediction of SHG response from more complex interfaces of all types, but also suggests new insights in the interpretation of the SHG signal at a molecular level. In particular, it highlights the modest influence of the molecular orientation and the high significance of the bulk quadrupole contribution, which does not depend on the interface, in the total experimental response.

| S-EPMC8612378 | biostudies-literature

Second harmonic generation imaging of corneal stroma after infection by Pseudomonas aeruginosa.

Project description:Pseudomonas aeruginosa is a pathogenic gram-negative organism that has the ability to cause blinding corneal infections following trauma and during contact lens wear. In this study, we investigated the directional movement and orientation of an invasive corneal isolate of P. aeruginosa in the corneal stroma during infection of ex vivo and in vivo rabbit corneas using multiphoton fluorescence and second harmonic generation (SHG) imaging. Ex vivo, rabbit corneas were subject to three partial thickness wounds prior to inoculation. In vivo, New Zealand white rabbits were fit with P. aeruginosa laden contact lenses in the absence of a penetrating wound. At all time points tested, infiltration of the corneal stroma by P. aeruginosa revealed a high degree of alignment between the bacteria and collagen lamellae ex vivo (p < 0.001). In vivo, P. aeruginosa traveled throughout the stroma in discrete regions or bands. Within each region, the bacteria showed good alignment with collagen lamellae (P = 0.002). Interestingly, in both the in vitro and in vivo models, P. aeruginosa did not appear to cross the corneal limbus. Taken together, our findings suggest that P. aeruginosa exploits the precise spacing of collagen lamellae in the central cornea to facilitate spread throughout the stroma.

| S-EPMC5387403 | biostudies-literature

Second-harmonic generation imaging of membrane potential with retinal analogues.

Project description:Second-harmonic generation (SHG) by membrane-incorporated probes is a nonlinear optical signal that is voltage-sensitive and the basis of a sensitive method for imaging membrane potential. The voltage dependence of SHG by four different probes, three retinoids (all-trans retinal), and two new retinal analogs, 3-methyl-7-(4'-dimethylamino-phenyl)-2,4,6-heptatrienal (AR-3) and 3,7-dimethyl-9-(4'-dimethylamino-phenyl)-2,4,6,8-nonatetraenal (AR-4), and a styryl dye (FM4-64), were compared in HEK-293 cells. Results were analyzed by fitting data with an expression based on an electrooptic mechanism for SHG, which depends on the complex-valued first- and second-order nonlinear electric susceptibilities (?² and ?³) of the probe. This gave values for the voltage sensitivity at the cell's resting potential, the voltage where the SHG is minimal, and the amplitude of the signal at that voltage for each of the four compounds. These measures show that ?² and ?³ are complex numbers for all compounds except all-trans retinal, consistent with the proximities of excitation and/or emission wavelengths to molecular resonances. Estimates of probe orientation and location in the membrane electric field show that, for the far-from-resonance case, the shot noise-limited signal/noise ratio depends on the location of the probe in the membrane, and on ?³ but not on ?².

| S-EPMC3297782 | biostudies-other

Chiral imaging of collagen by second-harmonic generation circular dichroism.

Project description:We provide evidence that the chirality of collagen can give rise to strong second-harmonic generation circular dichroism (SHG-CD) responses in nonlinear microscopy. Although chirality is an intrinsic structural property of collagen, most of the previous studies ignore that property. We demonstrate chiral imaging of individual collagen fibers by using a laser scanning microscope and type-I collagen from pig ligaments. 100% contrast level of SHG-CD is achieved with sub-micrometer spatial resolution. As a new contrast mechanism for imaging chiral structures in bio-tissues, this technique provides information about collagen morphology and three-dimensional orientation of collagen molecules.

| S-EPMC3675869 | biostudies-other

Second Harmonic Generation of Unpolarized Light.

Project description:A Mueller tensor mathematical framework was applied for predicting and interpreting the second harmonic generation (SHG) produced with an unpolarized fundamental beam. In deep tissue imaging through SHG and multiphoton fluorescence, partial or complete depolarization of the incident light complicates polarization analysis. The proposed framework has the distinct advantage of seamlessly merging the purely polarized theory based on the Jones or Cartesian susceptibility tensors with a more general Mueller tensor framework capable of handling partial depolarized fundamental and/or SHG produced. The predictions of the model are in excellent agreement with experimental measurements of z-cut quartz and mouse tail tendon obtained with polarized and depolarized incident light. The polarization-dependent SHG produced with unpolarized fundamental allowed determination of collagen fiber orientation in agreement with orthogonal methods based on image analysis. This method has the distinct advantage of being immune to birefringence or depolarization of the fundamental beam for structural analysis of tissues.

| S-EPMC5801736 | biostudies-literature

Second harmonic generation microscopy in zebrafish.

Project description:Modern optical imaging has progressed rapidly with the ability to noninvasively image cellular and subcellular phenomena with high spatial and temporal resolution. In particular, emerging techniques such as second harmonic generation (SHG) microscopy can allow for the monitoring of intrinsic contrast, such as that from collagen, in live and fixed samples. When coupled with multiphoton fluorescence microscopy, SHG can be used to image interactions between cells and the surrounding extracellular environment. There is recent interest in using these approaches to study inflammation and wound healing in zebrafish, an important model for studying these processes. In this chapter we present the practical aspects of using second harmonic generation to image interactions between leukocytes and collagen during wound healing in zebrafish.

| S-EPMC5489409 | biostudies-literature

New Details of the Human Corneal Limbus Revealed With Second Harmonic Generation Imaging.

Project description:PurposeTo report novel findings of the human corneal limbus by using second harmonic generation (SHG) imaging.MethodsCorneal limbus was imaged by using an inverted two-photon excitation fluorescence microscope. Laser (Ti:Sapphire) was tuned at 850 nm for two-photon excitation. Backscatter signals of SHG and autofluorescence (AF) were collected through a 425/30-nm emission filter and a 525/45-emission filter, respectively. Multiple, consecutive, and overlapping image stacks (z-stack) were acquired for the corneal limbal area.ResultsTwo novel collagen structures were revealed by SHG imaging at the limbus: an anterior limbal cribriform layer and presumed anchoring fibers. Anterior limbal cribriform layer is an intertwined reticular collagen architecture just beneath the limbal epithelial niche and is located between the peripheral cornea and Tenon's/scleral tissue. Autofluorescence imaging revealed high vascularity in this structure. Central to the anterior limbal cribriform layer, radial strands of collagen were found to connect the peripheral cornea to the limbus. These presumed anchoring fibers have both collagen and elastin and were found more extensively in the superficial layers than deep layer and were absent in very deep limbus near Schlemm's canal.ConclusionsBy using SHG imaging, new details of the collagen architecture of human corneal limbal area were elucidated. High resolution images with volumetric analysis revealed two novel collagen structures.

| S-EPMC4585533 | 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