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

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Nelly Vuillemin ◽  
Pierre Mahou ◽  
Delphine Débarre ◽  
Thierry Gacoin ◽  
Pierre-Louis Tharaux ◽  
...  
Keyword(s):  
Second Harmonic ◽  
Bessel Beam ◽  
Depth Of Field ◽  
Label Free ◽  
Focal Volume ◽  
Coherent Signals ◽  
Excitation Scheme ◽  
Collagen Accumulation ◽  
Excitation Volume ◽  
Beam Excitation

Abstract 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.

scholarly journals Erratum: Efficient second-harmonic imaging of collagen in histological slides using Bessel beam excitation

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Nelly Vuillemin ◽  
Pierre Mahou ◽  
Delphine Débarre ◽  
Thierry Gacoin ◽  
Pierre-Louis Tharaux ◽  
...  
Keyword(s):  
Second Harmonic ◽  
Bessel Beam ◽  
Harmonic Imaging ◽  
Beam Excitation

scholarly journals Advanced Label-Free Laser Scanning Microscopy and Its Biological Imaging Application

2021 ◽  
Vol 11 (3) ◽  
pp. 1002
Author(s):  
Xue Wang ◽  
Xinchao Lu ◽  
Chengjun Huang
Keyword(s):  
Laser Scanning ◽  
Second Harmonic ◽  
Biological Tissues ◽  
Biological Imaging ◽  
Laser Scanning Microscopy ◽  
Scanning Microscopy ◽  
Localized Surface Plasmon ◽  
Label Free ◽  
Coherent Raman ◽  
Surface Plasmon Microscopy

By eliminating the photodamage and photobleaching induced by high intensity laser and fluorescent molecular, the label-free laser scanning microscopy shows powerful capability for imaging and dynamic tracing to biological tissues and cells. In this review, three types of label-free laser scanning microscopies: laser scanning coherent Raman scattering microscopy, second harmonic generation microscopy and scanning localized surface plasmon microscopy are discussed with their fundamentals, features and recent progress. The applications of label-free biological imaging of these laser scanning microscopies are also introduced. Finally, the performance of the microscopies is compared and the limitation and perspectives are summarized.

Label-free second-harmonic phase imaging of biological specimen by digital holographic microscopy

2010 ◽  
Vol 35 (24) ◽  
pp. 4102 ◽  
Author(s):  
Etienne Shaffer ◽  
Corinne Moratal ◽  
Pierre Magistretti ◽  
Pierre Marquet ◽  
Christian Depeursinge
Keyword(s):  
Second Harmonic ◽  
Digital Holographic Microscopy ◽  
Phase Imaging ◽  
Label Free ◽  
Biological Specimen ◽  
Harmonic Phase ◽  
Holographic Microscopy ◽  
Harmonic Phase Imaging

Ultrahigh resolution optical coherence elastography using a Bessel beam for extended depth of field

2016 ◽  
Author(s):  
Andrea Curatolo ◽  
Martin Villiger ◽  
Dirk Lorenser ◽  
Philip Wijesinghe ◽  
Alexander Fritz ◽  
...  
Keyword(s):  
Bessel Beam ◽  
Depth Of Field ◽  
Optical Coherence ◽  
Ultrahigh Resolution ◽  
Extended Depth Of Field

Depth-of-field extension in optical imaging for rapid crystal screening

2021 ◽  
Vol 77 (4) ◽  
pp. 463-470
Author(s):  
Chen Li ◽  
Changqin Ding ◽  
Minghe Li ◽  
Jiayue Rong ◽  
Hilary Florian ◽  
...  
Keyword(s):  
Optical Imaging ◽  
Nonlinear Optical ◽  
Second Harmonic ◽  
Wavelength Dependence ◽  
Optical Microscope ◽  
Field Extension ◽  
Protein Crystal ◽  
Depth Of Field ◽  
Theoretical Predictions ◽  
Second Harmonic Generation Imaging

The depth of field (DoF) was extended 2.8-fold to achieve rapid crystal screening by retrofitting a custom-designed micro-retarder array (µRA) in the optical beam path of a nonlinear optical microscope. The merits of the proposed strategy for DoF enhancement were assessed in applications of second-harmonic generation imaging of protein crystals. It was found that DoF extension increased the number of crystals detected while simultaneously reducing the number of `z-slices' required for screening. Experimental measurements of the wavelength-dependence of the extended DoF were in excellent agreement with theoretical predictions. These results provide a simple and broadly applicable approach to increase the throughput of existing nonlinear optical imaging methods for protein crystal screening.

scholarly journals Multiscale molecular profiling of pathological bone resolves sexually dimorphic control of extracellular matrix composition

2021 ◽  
Vol 14 (3) ◽  
pp. dmm048116 ◽  
Author(s):  
Aikta Sharma ◽  
Alice Goring ◽  
Peter B. Johnson ◽  
Roger J. H. Emery ◽  
Eric Hesse ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Ex Vivo ◽  
Second Harmonic ◽  
Collagen Fibre ◽  
Matrix Composition ◽  
Label Free ◽  
Fibre Number ◽  
Backscattered Electron ◽  
Matrix Mineralisation

ABSTRACTCollagen assembly during development is essential for successful matrix mineralisation, which determines bone quality and mechanocompetence. However, the biochemical and structural perturbations that drive pathological skeletal collagen configuration remain unclear. Deletion of vascular endothelial growth factor (VEGF; also known as VEGFA) in bone-forming osteoblasts (OBs) induces sex-specific alterations in extracellular matrix (ECM) conformation and mineralisation coupled to vascular changes, which are augmented in males. Whether this phenotypic dimorphism arises as a result of the divergent control of ECM composition and its subsequent arrangement is unknown and is the focus of this study. Herein, we used murine osteocalcin-specific Vegf knockout (OcnVEGFKO) and performed ex vivo multiscale analysis at the tibiofibular junction of both sexes. Label-free and non-destructive polarisation-resolved second-harmonic generation (p-SHG) microscopy revealed a reduction in collagen fibre number in males following the loss of VEGF, complemented by observable defects in matrix organisation by backscattered electron scanning electron microscopy. This was accompanied by localised divergence in collagen orientation, determined by p-SHG anisotropy measurements, as a result of OcnVEGFKO. Raman spectroscopy confirmed that the effect on collagen was linked to molecular dimorphic VEGF effects on collagen-specific proline and hydroxyproline, and collagen intra-strand stability, in addition to matrix carbonation and mineralisation. Vegf deletion in male and female murine OB cultures in vitro further highlighted divergence in genes regulating local ECM structure, including Adamts2, Spp1, Mmp9 and Lama1. Our results demonstrate the utility of macromolecular imaging and spectroscopic modalities for the detection of collagen arrangement and ECM composition in pathological bone. Linking the sex-specific genetic regulators to matrix signatures could be important for treatment of dimorphic bone disorders that clinically manifest in pathological nano- and macro-level disorganisation.This article has an associated First Person interview with the first author of the paper.

scholarly journals Murine Metatarsus Bone and Joint Collagen-I Fiber Morphologies and Networks Studied With SHG Multiphoton Imaging

2021 ◽  
Vol 9 ◽  
Author(s):  
Martin Vielreicher ◽  
Aline Bozec ◽  
Georg Schett ◽  
Oliver Friedrich
Keyword(s):  
Rheumatoid Arthritis ◽  
Ex Vivo ◽  
Second Harmonic ◽  
Chronic Inflammatory Disease ◽  
Collagen I ◽  
Bone Architecture ◽  
Fiber Types ◽  
Label Free ◽  
Fiber Morphology ◽  
Collagen Network

Chronic inflammatory disease of bones and joints (e.g., rheumatoid arthritis, gout, etc.), but also acute bone injury and healing, or degenerative resorptive processes inducing osteoporosis, are associated with structural remodeling that ultimately have impact on function. For instance, bone stability is predominantly orchestrated by the structural arrangement of extracellular matrix fibrillar networks, i.e., collagen-I, -IV, elastin, and other proteins. These components may undergo distinct network density and orientation alterations that may be causative for decreased toughness, resilience and load bearing capacity or even increased brittleness. Diagnostic approaches are usually confined to coarse imaging modalities of X-ray or computer tomography that only provide limited optical resolution and lack specificity to visualize the fibrillary collagen network. However, studying collagen structure at the microscopic scale is of considerable interest to understand the mechanisms of tissue pathologies. Multiphoton Second Harmonic Generation (SHG) microscopy, is able to visualize the sterical topology of the collagen-I fibrillar network in 3D, in a minimally invasive and label-free manner. Penetration depths exceed those of conventional visible light imaging and can be further optimized through employing decalcification or optical clearing processing ex vivo. The goal of this proof-of-concept study was to use SHG and two-photon excited fluorescence (2-PEF) imaging to mainly characterize the fibrillary collagen organization within ex vivo decalcified normal mouse metatarsus bone and joint. The results show that the technique resolved the fibrillar collagen network of complete bones and joints with almost no artifacts and enabled to study the complex collagen-I networks with various fiber types (straight, crimped) and network arrangements of mature and woven bone with high degree of detail. Our imaging approach enabled to identify cavities within both cortical and trabecular bone architecture as well as interfaces with sharply changing fiber morphology and network structure both within bone, in tendon and ligament and within joint areas. These possibilities are highly advantageous since the technology can easily be applied to animal models, e.g., of rheumatoid arthritis to study structural effects of chronic joint inflammation, and to many others and to compare to the structure of human bone.

scholarly journals Generating Bessel beams with broad depth-of-field by using phase-only acoustic holograms

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Sergio Jiménez-Gambín ◽  
Noé Jiménez ◽  
José M. Benlloch ◽  
Francisco Camarena
Keyword(s):  
Field Distribution ◽  
Focal Length ◽  
Bessel Beam ◽  
Transverse Field ◽  
High Order ◽  
Depth Of Field ◽  
Uniform Field ◽  
Axial Distribution ◽  
Bessel Beams ◽  
Wide Range

AbstractWe report zero-th and high-order acoustic Bessel beams with broad depth-of-field generated using acoustic holograms. While the transverse field distribution of Bessel beams generated using traditional passive methods is correctly described by a Bessel function, these methods present a common drawback: the axial distribution of the field is not constant, as required for ideal Bessel beams. In this work, we experimentally, numerically and theoretically report acoustic truncated Bessel beams of flat-intensity along their axis in the ultrasound regime using phase-only holograms. In particular, the beams present a uniform field distribution showing an elongated focal length of about 40 wavelengths, while the transverse width of the beam remains smaller than 0.7 wavelengths. The proposed acoustic holograms were compared with 3D-printed fraxicons, a blazed version of axicons. The performance of both phase-only holograms and fraxicons is studied and we found that both lenses produce Bessel beams in a wide range of frequencies. In addition, high-order Bessel beam were generated. We report first order Bessel beams that show a clear phase dislocation along their axis and a vortex with single topological charge. The proposed method may have potential applications in ultrasonic imaging, biomedical ultrasound and particle manipulation applications using passive lenses.

Sign in / Sign up

Export Citation Format

Share Document