Opaque Tissue Research Articles

Ultrasound Biomicroscopy Diagnosis Analysis and Fine Care of Anterior Segment Injury of Traumatic Anterior Chamber Based on Intelligent Virtual Reality Technology.

The front part of the eye is exposed to the surface of the body, making it susceptible to injury. Blunt eye injuries often destroy the tissue structure in the anterior part of the eye. Often there is detachment of the iris root, separation of the ciliary body, the angle of retraction of the chamber, and complete dislocation or subluxation of the lens. Ordinary ocular ultrasound can penetrate into opaque tissue, but because of its low resolution, it can only show its outline, which is still very small for doctors in assisting diagnosis. Ultrasonic biomicroscopy is a very real-time analysis, will not be affected by the refraction space, and has great application value for ocular trauma. Based on this, this article proposes the ultrasound biomicroscope diagnosis analysis and fine nursing research of traumatic anterior chamber injury based on intelligent virtual reality technology. This article describes literature methods, experimental analysis methods, and other methods, in-depth study of the causes of traumatic anterior injuries, ultrasound biomicroscopy, and other theoretical knowledge, ultrasound biomicroscopy and microcontrol experiments for traumatic anterior injury design, and finally the causes of eye damage. This study compares the pairing results of ultrasound biomicroscopy biological measurements, analyzes the relationship between anterior bleeding level and anterior ocular injury, and analyzes the outcome and extent of service ophthalmology. The angle retreat accounted for 52.17% in grade I, 72.22% in grade II, and 60% in grade III. Anterior bleeding can cause single or complex iris root detachment, iris root detachment, ciliary body detachment, and angular leakage. The more the bleeding, the more serious the eye damage.

Topical Gadobutrol Application Causes Fluorescence Intensity Change in RFP-expressing Tumor-Bearing Mice

Optical clearing (OC) allows one to observe tissue structures and metabolic processes occurring in opaque tissues at the depths significantly exceeding the depths that can be reached without OC. Recently, we have shown that gadobutrol is a promising agent for OC of tissues in vivo. The aim of this study was to investigate the effect of time-dependence optical clearing caused by gadobutrol on intensity of fluorescent protein constitutively expressed in subcutaneous tumors in vivo. The measurements were performed in nu/nu mice bearing HEp-2 tumors expressing the red fluorescent protein TagRFP. Gadobutrol was used directly at concentrations 1.0 M aqueous solution or as a 0.7 M aqueous solution containing 5% dimethyl sulfoxide (DMSO). Gadobutrol was applied topically onto the skin above the tumors for 15 min. Tissue fluorescence was measured by using in vivo planar imaging technique. It was shown that the fluorescence intensity of tumors increased by 1.1–1.5 times in different animals under the influence of gadobutrol. The increase in intensity was more pronounced in the case of 0.7 M gadobutrol supplemented with DMSO. Apparently, the observed difference of penetration depths was due to the presence of DMSO in 0.7 M gadobutrol mixture.

High-resolution 3D fluorescent imaging of intact tissues.

Histological analysis of fluorescently labeled tissues has been a critical tool to understand molecular organization in situ. However, assessing molecular structures within large cells and in the context of human organ anatomy has been challenging because it requires penetration of staining reagents and light deep into opaque tissues, while also conforming to the spatial constraints of high-resolution objective lenses. This methodology article describes optimized sample preparation for sub-micron resolution 3D imaging in human and rodent tissues, yielding imaging depth (>100 μm) and resolution (<0.012 μm3 voxel size) that has previously been limited to whole-mount in vitro organoid systems, embryos, and small model organisms. Confocal images of adult human and rodent organs, including heart, kidney, and liver, were generated for several chemical and antibody stains in cleared tissue sections >100 μm thick. This method can be readily adopted by any lab performing routine histology and takes 3 days from the start of tissue preparation to 3D images.

Molecular Fluorophores for Deep-Tissue Bioimaging.

Fluorescence imaging has made tremendous inroads toward understanding the complexity of biological systems, but in vivo deep-tissue imaging remains a great challenge due to the optical opacity of biological tissue. Recent improvements in laser and detector manufacturing have allowed the expansion of nonlinear and linear fluorescence imaging to the underexplored “tissue-transparent” second near-infrared (NIR-II; 1000–1700 nm) window, opening up new opportunities for optical access deep inside opaque tissue. Molecular fluorophores have historically played a major role in fluorescence bioimaging. It is increasingly important to design new molecular fluorophores to fully unlock the potential of NIR-II imaging techniques. In this outlook, we give an overview of the novel molecular fluorophores developed for deep-tissue bioimaging in the past five years and discuss their pros and cons in applications. Guidelines for designing new molecular fluorophores with the desirable properties are also provided.

Photoacoustic Imaging to Track Magnetic-manipulated Micro-Robots in Deep Tissue.

The next generation of intelligent robotic systems has been envisioned as micro-scale mobile and externally controllable robots. Visualization of such small size microrobots to track their motion in nontransparent medium such as human tissue remains a major challenge, limiting translation into clinical applications. Herein, we present a novel, non-invasive, real-time imaging method by integrating ultrasound (US) and photoacoustic (PA) imaging modalities for tracking and detecting the motion of a single microrobot in deep biological tissue. We developed and evaluated a prototyped PA-guided magnetic microrobot tracking system. The microrobots are fabricated using photoresist mixed with nickel (Ni) particles. The microrobot motion was controlled using an externally applied magnetic field. Our experimental results evaluated the capabilities of PA imaging in visualizing and tracking microrobots in opaque tissue and tissue-mimicking phantoms. The results also demonstrate the ability of PA imaging in detecting a microrobot with the sizes less than the minimum detectable size by US imaging (down to 50 µm). The spectroscopic PA imaging studies determined an optimal wavelength (700 nm) for imaging microrobots with embedded Ni particles in oxygenated (fresh) human blood. In addition, we examined the ability of PA imaging to detect the microrobots through a nontransparent tissue mimic and at a depth of 25 mm, where conventional optical methods are unable to be used in tracking the objects. These initial results demonstrate the feasibility of an integrated US and PA imaging method to push the boundaries of microrobot applications into translational applications.

3D Imaging of Cleared Tissues

Tissue clearing technique enables visualization of opaque organ and tissue in 3‐dimension by turning tissue transparent. Current tissue clearing methods are restricted by limited types of tissue that can be cleared with each individual protocol, which inevitably led to the presence of blind‐spots within whole body or body parts imaging. Hard tissues including bones and teeth are still the most difficult organs to be cleared. In addition, endogenous fluorescence loss remains to be a major concern for solvent based clearing methods. Here, we developed a polyethylene glycol (PEG) Associated Solvent System (PEGASOS), which rendered nearly all types of tissue transparent and preserved endogenous fluorescence. Bones and teeth could be turned to nearly invisible after clearing. The PEGASOS method turned the whole adult mouse body transparent and we were able to image an adult mouse head composed of bones, teeth, brain, muscles and other tissues with no blind areas. Hard tissue transparency enabled us to reconstruct intact mandible, teeth, femur or knee joint in 3‐D. We were able to image intact mouse brain at sub‐cellular resolution and to trace individual neurons and axons over a long distance. We were able to visualize dorsal root ganglions directly through vertebrae. Finally, we revealed the neural network distribution pattern in 3‐dimension within the long bone marrow space. These results suggest that the PEGASOS method is a useful tool for general biomedical research.Support or Funding InformationThis work was supported by startup funding from Texas A&amp;M University, NIH/NIDCR grant K08DE025090, NIH/NIDCR grant R21 DE027928 and NIH/NIDCR grant R01 DE 028291 to Hu Zhao.Thy1‐EGFP spinal cord imaged with a tiling light sheet microscopeFigure 1

Measuring cell displacements in opaque tissues: dynamic light scattering in the multiple scattering regime.

Coherent light scattered by tissues brings structural and dynamic information, at depth, that standard imaging techniques cannot reach. Dynamics of cells or sub-cellular elements can be measured thanks to dynamic light scattering in thin samples (single scattering regime) or thanks to diffusive wave spectroscopy in thick samples (diffusion regime). Here, we address the intermediate regime and provide an analytical relationship between scattered light fluctuations and the distribution of cell displacements as a function of time. We illustrate our method by characterizing cell motility inside half millimeter thick multicellular aggregates.

Fibronectin has multifunctional roles in posterior capsular opacification (PCO)

Fibrotic posterior capsular opacification (PCO), one of the major complications of cataract surgery, occurs when lens epithelial cells (LCs) left behind post cataract surgery (PCS) undergo epithelial to mesenchymal transition, migrate into the optical axis and produce opaque scar tissue. LCs left behind PCS robustly produce fibronectin, although its roles in fibrotic PCO are not known. In order to determine the function of fibronectin in PCO pathogenesis, we created mice lacking the fibronectin gene (FN conditional knock out -FNcKO) from the lens. While animals from this line have normal lenses, upon lens fiber cell removal which models cataract surgery, FNcKO LCs exhibit a greatly attenuated fibrotic response from 3 days PCS onward as assessed by a reduction in surgery-induced cell proliferation, and fibrotic extracellular matrix (ECM) production and deposition. This is correlated with less upregulation of Transforming Growth Factor β (TGFβ) and integrin signaling in FNcKO LCs PCS concomitant with sustained Bone Morphogenetic Protein (BMP) signaling and elevation of the epithelial cell marker E cadherin. Although the initial fibrotic response of FNcKO LCs was qualitatively normal at 48 h PCS as measured by the upregulation of fibrotic marker protein αSMA, RNA sequencing revealed that the fibrotic response was already quantitatively attenuated at this time, as measured by the upregulation of mRNAs encoding molecules that control, and are controlled by, TGFβ signaling, including many known markers of fibrosis. Most notably, gremlin-1, a known regulator of TGFβ superfamily signaling, was upregulated sharply in WT LCs PCS, while this response was attenuated in FNcKO LCs. As exogenous administration of either active TGFβ1 or gremlin-1 to FNcKO lens capsular bags rescued the attenuated fibrotic response of fibronectin null LCs PCS including the loss of SMAD2/3 phosphorylation, this suggests that fibronectin plays multifunctional roles in fibrotic PCO development.

Sample Preparation of Optically Cleared Liver Tissue to Identify Liver Macrophages Using 3D Microscopy.

Histology, or the study of tissue microanatomy, is essential to understanding the in situ function of varying cell types within an organ. How cells are distributed throughout organs provides an indication of how they interact with other cells and structures within the organ microanatomy. The tortuous shape and large size of liver macrophages limits the value of standard tissue thickness of 5-10μm. As a result, imaging of specimens ideally thicker than 100μm is necessary to investigate the liver microanatomy and the how macrophages are distributed within this. Modern methods of microscopy, such as confocal and light sheet microscopy, allow for the analysis of tissue specimens of a thickness well beyond 100μm in the z-dimension. Liver tissue is an opaque tissue, and as a result, different techniques are needed to ameliorate light diffraction within the tissue. These techniques, in conjunction with antibody staining and refractive index matching of the tissue, have allowed researchers to image liver tissue specimens of more than 100μm thickness. Two of these techniques are modified versions of the clearing methods known as clearing-enhanced 3D (Ce3D) and fructose, urea, and glycerol for imaging (FUnGI). Here, we discuss the steps involved in preparing tissue specimens for optically clearing tissue using Ce3D and FUnGI for subsequent analysis of the distribution of macrophages in three dimensions using a confocal microscope.

A Novel Approach for Mammogram Enhancement using Nonlinear Unsharp Masking and L0 Gradient Minimization

Abstract Mammography is an effective modality for detecting and diagnosing breast cancer. Mammogram enhancement is used to improve the local contrast of mammograms, such that lesions appear more visible in the enhanced image. This paper introduces two novel approaches for mammogram enhancement based on a new unsharp masking scheme, called nonlinear UM and L0 Gradient Minimization, NLUML0GMIN for detecting breast cancer at an early stage. Three different techniques are unified in the proposed method; i) a Nonlinear filtering process to enhance the fine details within a 3x3 local neighborhood, ii) an L0 Gradient Minimization step globally preserving the high-contrast edges while suppressing low-contrast details seen as opaque fibrous tissues; and to obtain a detail mammogram after subtracting smoothed mammogram from the original mammogram, iii) Finally, an Unsharp masking technique combining detail mammogram with filtered mammogram passed through Non-linear filter, using PLIP operators which satisfy both Weber’s law and saturation characteristics of the human visual system.. An HVS-based decomposition scheme is used to analyse and visualize the malignant areas in the enhanced mammogram. The distinct arrangement of PLIP operators in the proposed framework has given rise to two methods NLUML0GMINAUTO and NLUML0GMIN, which outperforms NLUM method and other existing techniques for mammogram enhancement. The former is good at enhancing less-dense and medium-dense mammograms whereas the later enhances unseen masses within high-dense mammograms. Experimental results have stated the significance of the proposed method in effectively diagnosing breast cancer in comparison with the previous methods for mammogram enhancement. The mini-MIAS database has categorized mammograms as fatty, fatty-glandular and dense-glandular, which has helped us in focusing on enhancing images uniquely for each category. The proposed NLUML0GMIN scheme is robust and effective in identifying cancerous masses and calcifications within dense X-ray mammograms, aiding doctors in better cancer diagnosis by saving the life of countless patients with cancer.

A novel, adaptable, radiographically opaque, multi-plane continuous filament marker for optimizing tissue identification, radiation planning, and radiographic follow-up.

In breast cancer treatment, marking the tumor bed is an important aspect of the surgical component of therapy. Clear delineation of the tumor bed allows radiation oncologists a defined target for planning and delivering postoperative radiation therapy (XRT). Tumor bed marking also allows radiographic follow-up of the tumor bed on subsequent breast imaging. The aim of this assessment is to evaluate the ease and feasibility of utilizing a tumor bed filament marker (VeraFormÒ, Videra Surgical inc., USA) as a marker in post-operative benign surgical sites and malignant breast surgical tumor beds in breast cancer surgery. The filament marker is a novel radiopaque surgical filament that in lieu of clips and other markers is implanted in the surgical tumor bed during breast surgery. Following development of the filament marker, the researchers used breast phantoms and radiographic images to develop a series of geometric patterns of placement options that optimize comprehensive multi-plane radiographic interpretation of the exact tumor bed or surgical margin. Three breast surgeons at 3 separate institutions then used this filament as a continuous multi-plane marker in 20 patients during breast conservation surgery. In these patients, the filament marker was thus used to mark the tumor bed (breast cancer surgery) or surgical site (benign breast disease) instead of the more traditional devices such as clips or other metallic open framework devices. We then assessed 2 important factors related to this device; (I) the ease, feasibility, and accuracy of in vivo placement with oncoplastic and non-oncoplastic breast conservation surgery techniques; (II) the radiographic footprint this device left on standard imaging protocols of post-operative mammogram (MMG), computed tomography (CT) scan, breast magnetic resonance imaging (MRI) examinations, and ultrasounds (USs) for both routine follow-up imaging and for standard radiation planning. There were no adverse events reported with the use of this device. The cases were then reviewed by a multidisciplinary team that included the original surgeon, a breast radiologist, and radiation oncologist. Their unanimous evaluation was that the filament marker clearly delineated all sides and planes of the tumor bed (cancer surgery) or surgical site (benign disease). Regardless of surgical technique utilized, this information provided precise 3D guidance for radiation planning and delivery as well as radiographic follow-up. The surgeons involved reported that delineating the bed with the filament marker was a quick and easy procedure and did not interfere with performing the planned surgical technique. Radiologists, surgeons, and radiation oncologists found that the filament marker was not only radiographically opaque on CT and MMG, but also caused no significant artifact on CT, MRI, US, or MMG. The continuous multi-plane filament marker is a new device that fulfills the heretofore unmet need for safe and improved tumor bed and tissue site marking. It is an easy to place, non-palpable continuous multi-plane radiographic opaque tissue marker that seems to better delineate the tumor bed, regardless of type of breast surgery performed, while providing a more accurate 3D image for radiation planning and radiographic follow-up on MMG MRI, CT and US.

Corneal Transplantation Today. Giving Patients A Better Quality of Life

Corneal transplant has been around for many years, the first one done more than a 100 years ago. Although the principle is the same, exchanging unhealthy, distorted, opaque or decompensated tissue for a healthy one, thanks to the evolution, knowledge in Medicine, higher technology and the need for more successful results, corneal transplant has reached a very specialized status.

AC–DC Electropenetrography for In-depth Studies of Feeding and Oviposition Behaviors

Studying the feeding, host damage, and transmission (i.e., acquisition, retention, and inoculation) of pathogens (plant and animal) by piercing-sucking arthropods is challenging. The piercing mouthparts are probed into opaque host tissues, precluding direct observation. This challenge was overcome by the invention of electropenetrography, or electrical penetration graph (both abbreviated EPG), the most rigorous method to identify arthropod feeding behaviors. However, until recently, most EPG research was restricted to studies of hemipteran plant pests, especially aphids and leafhoppers. Recent advances in EPG technology via the third-generation, alternating current–direct current (AC–DC) monitor (electropenetrograph) are opening new doors to expand the science into all types of arthropod feeding and oviposition. EPG can be used in three ways for the development of novel integrated pest management (IPM) tactics. First, in cases where the fundamental mechanisms of feeding/oviposition effects or pathogen transmission are unknown, EPG is instrumental in identifying mechanisms. Second, once the causes of damage or transmission are understood, EPG can be used to demonstrate the effects of insecticides, antifeedants, repellents, or other chemicals on specific feeding behaviors responsible for damage or transmission.Third, EPG can similarly identify the effects of resistant versus susceptible varieties of crop plants and animals, including transgenic organisms genetically engineered to express biopesticides or other genes. The purpose of this paper is to encourage new research avenues by 1) thoroughly reviewing history and electronic principles of EPG, culminating in the AC–DC electropenetrograph, 2) reviewing principles underlying biological meanings of waveforms, and 3) presenting a few examples of waveforms from new arthropod taxa.

A Novel Optical Tissue Clearing Protocol for Mouse Skeletal Muscle to Visualize Endplates in Their Tissue Context

Neuromuscular junctions (NMJs) mediate skeletal muscle contractions and play an important role in several neuromuscular disorders when their morphology and function are compromised. However, due to their small size and sparse distribution throughout the comparatively large, inherently opaque muscle tissue the analysis of NMJ morphology has been limited to teased fiber preparations, longitudinal muscle sections, and flat muscles. Consequently, whole mount analyses of NMJ morphology, numbers, their distribution, and assignment to a given muscle fiber have also been impossible to determine in muscle types that are frequently used in experimental paradigms. This impossibility is exacerbated by the lack of optical tissue clearing techniques that are compatible with clear and persistent NMJ stains. Here, we present MYOCLEAR, a novel and highly reproducible muscle tissue clearing protocol. Based on hydrogel-based tissue clearing methods, this protocol permits the labeling and detection of all NMJs in adult hindleg extensor digitorum longus muscles from wildtype and diseased mice. The method is also applicable to adult mouse diaphragm muscles and can be used for different staining agents, including toxins, lectins, antibodies, and nuclear dyes. It will be useful in understanding the distribution, morphological features, and muscle tissue context of NMJs in hindleg muscle whole mounts for biomedical and basic research.

Immunoresolvent Resolvin D1 Maintains the Health of the Ocular Surface.

The present review focuses on the role of one of the D-series resolvins (Rv) RvD1 in the regulation of conjunctival goblet cell secretion and its role in ocular surface health. RvD1 is the most thoroughly studied of the specialized proresolution mediators in the goblet cells. The anterior surface of the eye consists of the cornea (the transparent central area) and the conjunctiva (opaque tissue that surrounds the cornea and lines the eyelids). The secretory mucin MUC5AC produced by the conjunctival goblet cells is protective of the ocular surface and especially helps to maintain clear vision through the cornea. In health, a complex neural reflex stimulates goblet cell secretion to maintain an optimum amount of mucin in the tear film. The specialized pro-resolution mediator, D-series resolvin (RvD1) is present in human tears and induces goblet cell mucin secretion. RvD1 interacts with its receptors ALX/FPR2 and GPR32, activates phospholipases C, D, and A2, as well as the EGFR. This stimulation increases the intracellular [Ca2+] and activates extracellular regulated kinase (ERK) 1/2 to cause mucin secretion into the tear film. This mucin secretion protects the ocular surface from the challenges in the external milieu thus maintaining a healthy interface between the eye and the environment. RvD1 forms a second important mechanism along with activation of a neural reflex pathway to regulate goblet cell mucin secretion and protect the ocular surface in health.

A method for transplantation of human HSCs into zebrafish, to replace humanised murine transplantation models.

Haematopoietic stem cell (HSC) transplantation is a critical therapy for haematopoietic malignancies and immune disorders. Incomplete or delayed engraftment of HSCs in the host results in increased risk of infection and morbidity. The mechanisms of HSC engraftment are poorly understood and understanding these processes will increase transplantation success on many levels. Current animal models are immunocompromised 'humanised' mice transplanted with human HSCs. Harmful procedures include genetic manipulations and irradiation to ablate the mouse immune system, and opaque mouse tissues make visualisation of the early steps of HSC engraftment impossible. There is a need for new models to offer alternatives to humanised mice in the study of HSC transplantation. Here we described a detailed method for transplantation of human HSCs into zebrafish, before the onset of adaptive immunity. Human HSCs were purified from whole blood by enrichment of the CD34 cell population using a positive magnetic selection and further purified using an anti-CD34 antibody and cell sorting. Sorted CD34 cells were transplanted into the blood stream of 52 hour old zebrafish larvae. Human HSCs home into the zebrafish haematopoietic niche, where they engage with endothelial cells and undergo cell division. Our model offers the opportunities to image in vivo human HSC engraftment in a transparent organism, without the myeloablative strategies used in mice, and provides a unique system to understand the dynamic process of engraftment and replace current murine models. This technique can be applied to current engraftment protocols to validate the viability and efficiency of cryofrozen HSC grafts. This humanised zebrafish model will be instrumental to develop the 3Rs values in stem cell transplantation research and our detailed protocol will increase the chances of uptake of this zebrafish model by the mouse community.

Phase contrast MRI of creeping flows using stimulated echo.

Creeping flows govern many important physiological phenomena such as elevated interstitial fluid flows in tumors, glymphatic flows in the brain, among other applications. However, few methods exist to measure such slow flows non-invasively in optically opaque biological tissues in vivo. Phase-contrast MRI is a velocimetry technique routinely used in the clinic to measure fast flows in biological tissues, such as blood and cerebrospinal fluid (CSF), in the order of cm/s. Use of this technique to encode slower flows is hampered by diffusion weighting and phase error introduced by gradient hardware imperfections. In this study, a new PC-MRI technique is developed using stimulated echo preparation to overcome these challenges. Flows as slow as 1 μm/s are measured and validated using controlled water flow through a pipe at 4.7 T. The error in measured flow rate obtained by integrating the measured velocity over the cross-sectional area of the pipe is less than 10%. The developed method was also able to capture slow natural convection flows appearing in liquids placed inside a horizontal bore magnet. Monitoring the 4D velocity vector field revealed that the natural convection flows decay exponentially with time. This method could be applied in future to study creeping flows, e.g. in tissue.

Tomographic analysis of siliceous particulates in Australian turpentine wood (Syncarpia glomulifera) through X-ray micro-computed tomography

X-ray micro-computed tomography (XμCT) allows a non-destructive and three-dimensional (3D) study of otherwise complex and opaque wood tissues. In wood research, XμCT datasets are highly useful for the qualitative and quantitative examination of wood structures. In this study, XμCT was introduced and tested for examining X-ray dense silica particles in the Australian turpentine wood (Syncarpia glomulifera). It was possible to three-dimensionally visualize and numerically quantify silica particles. Numerical analysis was performed to scrutinize the size and content of silica particles. In comparative studies of silica size through scanning electron microscopy and silica content through thermo-gravimetric analysis after acid digestion of ash, our findings pointed out that XμCT is indeed a powerful tool for examining silica particles in wood; because XμCT enables a simultaneous visualization and quantification of the silica particles in 3D without being destructive. Despite these benefits, comparative examination through scanning electron microscopy and energy-dispersive X-ray spectroscopy is necessary to verify silica particles in tomographic images. XμCT technology might further aid in probing the biological and ecological function of silica in silica-bearing wood species.

Visualization of Schlemm's canal from 3D reconstruction of UMB images

Ultrasound biomicroscopy (UBM) is widely used by ophthalmologists for visualizing anterior segment structures. Compared with other ophthalmic imaging modalities, UBM excels in its ability to penetrate opaque tissue and visualize structures such as Schlemm's canal which cannot be seen through the lens.

Tissue clearing of both hard and soft tissue organs with the PEGASOS method

Tissue clearing technique enables visualization of opaque organs and tissues in 3-dimensions (3-D) by turning tissue transparent. Current tissue clearing methods are restricted by limited types of tissues that can be cleared with each individual protocol, which inevitably led to the presence of blind-spots within whole body or body parts imaging. Hard tissues including bones and teeth are still the most difficult organs to be cleared. In addition, loss of endogenous fluorescence remains a major concern for solvent-based clearing methods. Here, we developed a polyethylene glycol (PEG)-associated solvent system (PEGASOS), which rendered nearly all types of tissues transparent and preserved endogenous fluorescence. Bones and teeth could be turned nearly invisible after clearing. The PEGASOS method turned the whole adult mouse body transparent and we were able to image an adult mouse head composed of bones, teeth, brain, muscles, and other tissues with no blind areas. Hard tissue transparency enabled us to reconstruct intact mandible, teeth, femur, or knee joint in 3-D. In addition, we managed to image intact mouse brain at sub-cellular resolution and to trace individual neurons and axons over a long distance. We also visualized dorsal root ganglions directly through vertebrae. Finally, we revealed the distribution pattern of neural network in 3-D within the marrow space of long bone. These results suggest that the PEGASOS method is a useful tool for general biomedical research.