Low-cost, chromatic confocal endomicroscope for cellular imaging in vivo

The sensory end organ of the ear, the organ of Corti, rests on a thin basilar membrane which lies between the bone of the central modiolus and the bony wall of the cochlea. In vivo, the organ of Corti is protected by the bony wall which totally surrounds it. In order to examine the sensory epithelium by scanning electron microscopy it is necessary to dissect away the protective bone and expose the region of interest (Fig. 1). This leaves the fragile organ of Corti susceptible to physical damage during subsequent handling. In our laboratory cochlear specimens, after dissection, are routinely prepared by the O-T- O-T-O technique, critical point dried and then lightly sputter coated with gold. This processing involves considerable specimen handling including several hours on a rotator during which the organ of Corti is at risk of being physically damaged. The following procedure uses low cost, readily available materials to hold the specimen during processing ,preventing physical damage while allowing an unhindered exchange of fluids.Following fixation, the cochlea is dehydrated to 70% ethanol then dissected under ethanol to prevent air drying. The holder is prepared by punching a hole in the flexible snap cap of a Wheaton vial with a paper hole punch. A small amount of two component epoxy putty is well mixed then pushed through the hole in the cap. The putty on the inner cap is formed into a “cup” to hold the specimen (Fig. 2), the putty on the outside is smoothed into a “button” to give good attachment even when the cap is flexed during handling (Fig. 3). The cap is submerged in the 70% ethanol, the bone at the base of the cochlea is seated into the cup and the sides of the cup squeezed with forceps to grip it (Fig.4). Several types of epoxy putty have been tried, most are either soluble in ethanol to some degree or do not set in ethanol. The only putty we find successful is “DUROtm MASTERMENDtm Epoxy Extra Strength Ribbon” (Loctite Corp., Cleveland, Ohio), this is a blue and yellow ribbon which is kneaded to form a green putty, it is available at many hardware stores.

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Quantum Dot Nanocrystals for In Vivo Molecular and Cellular Imaging¶

Photochemistry and Photobiology ◽

10.1562/2004-06-21-ir-209.1 ◽

2004 ◽

Vol 80 (3) ◽

pp. 377 ◽

Cited By ~ 119

Author(s):

Andrew M. Smith ◽

Xiaohu Gao ◽

Shuming Nie

Keyword(s):

Quantum Dot ◽

Cellular Imaging

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Organ-specific, multimodal, wireless optoelectronics for high-throughput phenotyping of peripheral neural pathways

Nature Communications ◽

10.1038/s41467-020-20421-8 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Woo Seok Kim ◽

Sungcheol Hong ◽

Milenka Gamero ◽

Vivekanand Jeevakumar ◽

Clay M. Smithhart ◽

...

Keyword(s):

High Throughput ◽

Low Cost ◽

Optoelectronic Device ◽

Antenna System ◽

Neural Pathways ◽

Organ Specific ◽

Coil Antenna ◽

Sensory Fibers

AbstractThe vagus nerve supports diverse autonomic functions and behaviors important for health and survival. To understand how specific components of the vagus contribute to behaviors and long-term physiological effects, it is critical to modulate their activity with anatomical specificity in awake, freely behaving conditions using reliable methods. Here, we introduce an organ-specific scalable, multimodal, wireless optoelectronic device for precise and chronic optogenetic manipulations in vivo. When combined with an advanced, coil-antenna system and a multiplexing strategy for powering 8 individual homecages using a single RF transmitter, the proposed wireless telemetry enables low cost, high-throughput, and precise functional mapping of peripheral neural circuits, including long-term behavioral and physiological measurements. Deployment of these technologies reveals an unexpected role for stomach, non-stretch vagal sensory fibers in suppressing appetite and demonstrates the durability of the miniature wireless device inside harsh gastric conditions.

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Synthesis of Exosome-Based Fluorescent Gold Nanoclusters for Cellular Imaging Applications

International Journal of Molecular Sciences ◽

10.3390/ijms22094433 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4433

Author(s):

Eun Sung Lee ◽

Byung Seok Cha ◽

Seokjoon Kim ◽

Ki Soo Park

Keyword(s):

Photoelectron Spectroscopy ◽

Low Cost ◽

Stokes Shift ◽

Gold Nanoclusters ◽

Human Colon ◽

Cellular Imaging ◽

Metal Nanoclusters ◽

Human Colon Cancer ◽

X Ray ◽

High Concentrations

In recent years, fluorescent metal nanoclusters have been used to develop bioimaging and sensing technology. Notably, protein-templated fluorescent gold nanoclusters (AuNCs) are attracting interest due to their excellent fluorescence properties and biocompatibility. Herein, we used an exosome template to synthesize AuNCs in an eco-friendly manner that required neither harsh conditions nor toxic chemicals. Specifically, we used a neutral (pH 7) and alkaline (pH 11.5) pH to synthesize two different exosome-based AuNCs (exo-AuNCs) with independent blue and red emission. Using field-emission scanning electron microscopy, energy dispersive X-ray microanalysis, nanoparticle tracking analysis, and X-ray photoelectron spectroscopy, we demonstrated that AuNCs were successfully formed in the exosomes. Red-emitting exo-AuNCs were found to have a larger Stokes shift and a stronger fluorescence intensity than the blue-emitting exo-AuNCs. Both exo-AuNCs were compatible with MCF-7 (human breast cancer), HeLa (human cervical cancer), and HT29 (human colon cancer) cells, although blue-emitting exo-AuNCs were cytotoxic at high concentrations (≥5 mg/mL). Red-emitting exo-AuNCs successfully stained the nucleus and were compatible with membrane-staining dyes. This is the first study to use exosomes to synthesize fluorescent nanomaterials for cellular imaging applications. As exosomes are naturally produced via secretion from almost all types of cell, the proposed method could serve as a strategy for low-cost production of versatile nanomaterials.

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Optimizing the Reaction Conditions for the Formation of Fumarate via Trans-Hydrogenation

Applied Magnetic Resonance ◽

10.1007/s00723-021-01371-w ◽

2021 ◽

Author(s):

Laura Wienands ◽

Franziska Theiß ◽

James Eills ◽

Lorenz Rösler ◽

Stephan Knecht ◽

...

Keyword(s):

Aqueous Solution ◽

Thermal Equilibrium ◽

Reaction Rate ◽

Low Cost ◽

Hydrogen Gas ◽

Metabolic Imaging ◽

Reaction Conditions ◽

Precursor Molecule ◽

Solution Preparation

AbstractParahydrogen-induced polarization is a hyperpolarization method for enhancing nuclear magnetic resonance signals by chemical reactions/interactions involving the para spin isomer of hydrogen gas. This method has allowed for biomolecules to be hyperpolarized to such a level that they can be used for real time in vivo metabolic imaging. One particularly promising example is fumarate, which can be rapidly and efficiently hyperpolarized at low cost by hydrogenating an acetylene dicarboxylate precursor molecule using parahydrogen. The reaction is relatively slow compared to the timescale on which the hyperpolarization relaxes back to thermal equilibrium, and an undesirable 2nd hydrogenation step can convert the fumarate into succinate. To date, the hydrogenation chemistry has not been thoroughly investigated, so previous work has been inconsistent in the chosen reaction conditions in the search for ever-higher reaction rate and yield. In this work we investigate the solution preparation protocols and the reaction conditions on the rate and yield of fumarate formation. We report conditions to reproducibly yield over 100 mM fumarate on a short timescale, and discuss aspects of the protocol that hinder the formation of fumarate or lead to irreproducible results. We also provide experimental procedures and recommendations for performing reproducible kinetics experiments in which hydrogen gas is repeatedly bubbled into an aqueous solution, overcoming challenges related to the viscosity and surface tension of the water.

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Genome-wide integration site detection using Cas9 enriched amplification-free long-range sequencing

Nucleic Acids Research ◽

10.1093/nar/gkaa1152 ◽

2020 ◽

Author(s):

Joost van Haasteren ◽

Altar M Munis ◽

Deborah R Gill ◽

Stephen C Hyde

Keyword(s):

Long Range ◽

Insertional Mutagenesis ◽

Lentiviral Vector ◽

Integration Site ◽

Low Cost ◽

Safety Evaluation ◽

Read Length ◽

Chromosomal Dna ◽

Wide Range

Abstract The gene and cell therapy fields are advancing rapidly, with a potential to treat and cure a wide range of diseases, and lentivirus-based gene transfer agents are the vector of choice for many investigators. Early cases of insertional mutagenesis caused by gammaretroviral vectors highlighted that integration site (IS) analysis was a major safety and quality control checkpoint for lentiviral applications. The methods established to detect lentiviral integrations using next-generation sequencing (NGS) are limited by short read length, inadvertent PCR bias, low yield, or lengthy protocols. Here, we describe a new method to sequence IS using Amplification-free Integration Site sequencing (AFIS-Seq). AFIS-Seq is based on amplification-free, Cas9-mediated enrichment of high-molecular-weight chromosomal DNA suitable for long-range Nanopore MinION sequencing. This accessible and low-cost approach generates long reads enabling IS mapping with high certainty within a single day. We demonstrate proof-of-concept by mapping IS of lentiviral vectors in a variety of cell models and report up to 1600-fold enrichment of the signal. This method can be further extended to sequencing of Cas9-mediated integration of genes and to in vivo analysis of IS. AFIS-Seq uses long-read sequencing to facilitate safety evaluation of preclinical lentiviral vector gene therapies by providing IS analysis with improved confidence.

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Hydroxyapatite-incorporation improves bone formation on endosseous PEEK implant in canine tibia

Journal of Applied Biomaterials & Functional Materials ◽

10.1177/2280800020975172 ◽

2020 ◽

Vol 18 ◽

pp. 228080002097517

Author(s):

Yuan-ming Geng ◽

Dong-ni Ren ◽

Shu-yi Li ◽

Zong-yi Li ◽

Xiao-qing Shen ◽

...

Keyword(s):

Bone Formation ◽

Dental Implants ◽

Low Cost ◽

Low Elastic Modulus ◽

Ether Ether Ketone ◽

Poly Ether Ether Ketone ◽

Alternative Material ◽

Ct Analysis ◽

Potential Alternative

Background: Poly Ether Ether Ketone (PEEK) has been considered as a potential alternative material for endosseous dental implants, for its low elastic modulus, biocompatibility, and low cost in customized device manufacture. Hydroxyapatite-incorporation is supposed to improve the poor osseointegration of PEEK. Methods: In the present study we analyzed the in vivo response of hydroxyapatite-incorporated PEEK (PEEK-HA) implants in canine tibia. PEEK-HA and PEEK implants were implanted and were examined 4 weeks and 12 weeks after implantation with radiology and histology. Commercial titanium dental implants served as controls. Results: The ratio of bone volume to tissue volume of PEEK-HA implants was higher than that of PEEK implants 4 weeks after implantation in the μ-CT analysis. The bone implant contact of PEEK and PEEK-HA implants showed no statistical difference in the histological examination, but newly-formed bone around PEEK-HA implants showed more signs of mineralization than that around PEEK implants. Conclusion: The study suggested that bone formation was improved with hydroxyapatite-incorporation in PEEK. Hydroxyapatite-incorporated PEEK implants may represent a potential material for endosseous dental implant.

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Proanthocyanidins against Oxidative Stress: From Molecular Mechanisms to Clinical Applications

BioMed Research International ◽

10.1155/2018/8584136 ◽

2018 ◽

Vol 2018 ◽

pp. 1-11 ◽

Cited By ~ 12

Author(s):

Lingyu Yang ◽

Dehai Xian ◽

Xia Xiong ◽

Rui Lai ◽

Jing Song ◽

...

Keyword(s):

Oxidative Stress ◽

Signaling Pathways ◽

Molecular Mechanisms ◽

Metabolic Diseases ◽

Low Cost ◽

Natural Agents ◽

Molecular Damage ◽

And Control

Proanthocyanidins (PCs) are naturally occurring polyphenolic compounds abundant in many vegetables, plant skins (rind/bark), seeds, flowers, fruits, and nuts. Numerousin vitroandin vivostudies have demonstrated myriad effects potentially beneficial to human health, such as antioxidation, anti-inflammation, immunomodulation, DNA repair, and antitumor activity. Accumulation of prooxidants such as reactive oxygen species (ROS) exceeding cellular antioxidant capacity results in oxidative stress (OS), which can damage macromolecules (DNA, lipids, and proteins), organelles (membranes and mitochondria), and whole tissues. OS is implicated in the pathogenesis and exacerbation of many cardiovascular, neurodegenerative, dermatological, and metabolic diseases, both through direct molecular damage and secondary activation of stress-associated signaling pathways. PCs are promising natural agents to safely prevent acute damage and control chronic diseases at relatively low cost. In this review, we summarize the molecules and signaling pathways involved in OS and the corresponding therapeutic mechanisms of PCs.

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Compact, low-cost Ti:Al_2O_3 laser for in vivo ultrahigh-resolution optical coherence tomography

Optics Letters ◽

10.1364/ol.28.000905 ◽

2003 ◽

Vol 28 (11) ◽

pp. 905 ◽

Cited By ~ 58

Author(s):

A. Unterhuber ◽

B. Považay ◽

B. Hermann ◽

H. Sattmann ◽

W. Drexler ◽

...

Keyword(s):

Optical Coherence Tomography ◽

Low Cost ◽

Optical Coherence ◽

Ultrahigh Resolution

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