Accedi

NIH

14 ARTICLES PUBLISHED IN JoVE

image

Neuroscience

DiOLISTIC Labeling of Neurons from Rodent and Non-human Primate Brain Slices
Gail K. Seabold 1, James B. Daunais 2, Andrew Rau 3, Kathleen A. Grant 3, Veronica A. Alvarez 1
1Section on Neuronal Structure, Laboratory for Integrative Neuroscience, NIAAA, NIH, 2Department Physiology and Pharmacology, Wake Forest University Health Sciences, 3Oregon National Primate Research Center, Division of Neuroscience, Oregon Health and Science University

We demonstrate the use of the gene gun to introduce fluorescent dyes, such as DiI, into neurons in brain slices from rodents and non-human primates of different ages. In this particular case, we use adult mice (3-6 months old) and adult cynomologus monkeys (9-15 years old). This technique, originally described by the laboratory of Dr. Lichtman (Gan et al., 2000), is well suited for the study of dendritic branching and dendritic spine morphology and can be combined with traditional immunostaining, if detergents are kept at a low concentration.

image

Neuroscience

An Optic Nerve Crush Injury Murine Model to Study Retinal Ganglion Cell Survival
Zhongshu Tang 1, Shuihua Zhang 1,2, Chunsik Lee 1, Anil Kumar 1, Pachiappan Arjunan 1, Yang Li 1, Fan Zhang 1, Xuri Li 1
1National Eye Institute, NIH, 2Ophthalmology Department, The Second Hospital of Harbin Medical University

This protocol shows how to retrogradely label retinal ganglion cells, and how to subsequently make an optic nerve crush injury in order to analyze retinal ganglion cell survival and apoptosis. It is an experimental disease model for different types of optic neuropathy, including glaucoma.

image

Medicine

A Mouse Model of the Cornea Pocket Assay for Angiogenesis Study
Zhongshu Tang 1, Fan Zhang 1, Yang Li 1, Pachiappan Arjunan 1, Anil Kumar 1, Chunsik Lee 1, Xuri Li 1
1National Eye Institute

The cornea is unique in that it lacks vascular tissues. However, robust blood vessel growth and survival can be induced in the cornea by potent angiogenic factors. Therefore, the cornea can provide with us a valuable tool for angiogenic studies. This protocol demonstrates how to perform the mouse model of cornea pocket assay and how to assess the angiogenesis induced by angiogenic factors using this model.

image

Bioengineering

Imaging Denatured Collagen Strands In vivo and Ex vivo via Photo-triggered Hybridization of Caged Collagen Mimetic Peptides
Yang Li 1, Catherine A. Foss 2, Martin G. Pomper 2,3, S. Michael Yu 1,3
1Department of Bioengineering, University of Utah, 2Department of Radiology and Radiological Science, Johns Hopkins University School of Medicine, 3Institute for NanoBiotechnology, Johns Hopkins University

This procedure demonstrates in vivo near IR fluorescence imaging of collagen remodeling activities in mice as well as ex vivo staining of collagens in tissue sections using caged collagen mimetic peptides that can be photo-triggered to hybridize with denatured collagen strands.

image

Bioengineering

Engineering Platform and Experimental Protocol for Design and Evaluation of a Neurally-controlled Powered Transfemoral Prosthesis
Fan Zhang 1, Ming Liu 1, Stephen Harper 2,3, Michael Lee 3, He Huang 1
1Joint Department of Biomedical Engineering, North Carolina State University & University of North Carolina at Chapel Hill, 2Department of Physical Medicine and Rehabilitation, University of North Carolina School of Medicine, 3Atlantic Prosthetics & Orthotics, LLC

Neural-machine interfaces (NMI) have been developed to identify the user's locomotion mode. These NMIs are potentially useful for neural control of powered artificial legs, but have not been fully demonstrated. This paper presented (1) our designed engineering platform for easy implementation and development of neural control for powered lower limb prostheses and (2) an experimental setup and protocol in a laboratory environment to evaluate neurally-controlled artificial legs on patients with lower limb amputations safely and efficiently.

image

Immunology and Infection

Isolation, Characterization and Functional Examination of the Gingival Immune Cell Network
Nicolas Dutzan *1, Loreto Abusleme *1, Joanne E. Konkel 2,3, Niki M. Moutsopoulos 1
1Oral Immunity and Inflammation Unit, NIDCR, NIH, 2Manchester Immunology Group, Faculty of Life Sciences, University of Manchester, 3Manchester Collaborative Centre for Inflammation Research, University of Manchester

We have established a technique for the isolation, phenotypic characterization and functional analysis of immune cells from murine gingiva.

image

Immunology and Infection

An In Vitro Caseum Binding Assay that Predicts Drug Penetration in Tuberculosis Lesions
Jansy P. Sarathy 1, Hsin-pin Ho Liang 1, Danielle Weiner 2, Jacqueline Gonzales 2, Laura E. Via 2, Véronique Dartois 1
1Public Health Research Institute Centre, New Jersey Medical School, Rutgers, 2Tuberculosis Research Section, Laboratory of Clinical Infectious Diseases, NIAID, NIH

Here we describe a rapid equilibrium dialysis (RED) method to measure drug binding to caseum from pulmonary tuberculosis lesions and cavities. The protocol is also used with a foamy macrophage-derived matrix that is an effective surrogate to caseum.

image

Immunology and Infection

Single-cell Quantitation of mRNA and Surface Protein Expression in Simian Immunodeficiency Virus-infected CD4+ T Cells Isolated from Rhesus macaques
Andrey Tokarev 1, Matthew Creegan 1, Michael A. Eller 1, Mario Roederer 2, Diane L. Bolton 1
1US Military HIV Research Program, Henry M. Jackson Foundation, Walter Reed Army Institute of Research, 2Vaccine Research Center, NIAID, NIH

Described is a methodology to quantitate the expression of 96 genes and 18 surface proteins by single cells ex vivo, allowing for the identification of differentially expressed genes and proteins in virus-infected cells relative to uninfected cells. We apply the approach to study SIV-infected CD4+ T cells isolated from rhesus macaques.

image

Neuroscience

In Vitro Wedge Slice Preparation for Mimicking In Vivo Neuronal Circuit Connectivity
Matthew J. Fischl 1, Catherine J. C. Weisz 1
1Section on Neuronal Circuitry, National Institute on Deafness and Other Communication Disorders, NIH

Integration of diverse synaptic inputs to neurons is best measured in a preparation that preserves all pre-synaptic nuclei for natural timing and circuit plasticity, but brain slices typically sever many connections. We developed a modified brain slice to mimic in vivo circuit activity while maintaining in vitro experimentation capability.

image

Immunology and Infection

User-friendly, High-throughput, and Fully Automated Data Acquisition Software for Single-particle Cryo-electron Microscopy
Anil Kumar 1, Surekha P. 1, Sahil Gulati 2, Somnath Dutta 1
1Molecular Biophysics Unit, Indian Institute of Science, 2Gatan Inc.

Single-particle cryo-electron microscopy demands a suitable software package and user-friendly pipeline for high-throughput automatic data acquisition. Here, we present the application of a fully automated image acquisition software package, Latitude-S, and a practical pipeline for data collection of vitrified biomolecules under low-dose conditions.

image

Medicine

Isolation of Monocyte-Macrophage Lineage Cells from Rat Bones by Secondary Adherence Method
Xiaoli Jin *1, Yang Li *2, Xuanwei Chen 1, Jin Chen 1, Jian Xu 1
1School of Medical Technology and Information Engineering, Zhejiang Chinese Medical University, 2School of Basic Medical Sciences, Fudan University

Here we present a protocol for the isolation of BMMs from SD rats, called the secondary adherence method.

image

Cancer Research

Co-Culture and Transduction of Murine Thymocytes on Delta-Like 4-Expressing Stromal Cells to Study Oncogenes in T-Cell Leukemia
Gisele O. L. Rodrigues 1, WenQing Li 1, Sarah D. Cramer 1,2,3, Hila Y. Winer 1, Tu Chun Hsu 1,2,3, Timothy Gower 4, Julie A. Hixon 1, Scott K. Durum 1
1Cytokines and Immunity Section, Cancer Innovation Laboratory, National Cancer Institute, National Institutes of Health, 2Comparative Biomedical Scientist Training Program, NIH, 3Department of Pathobiology and Diagnostic Investigation, Veterinary Diagnostic Laboratory, Michigan State University, 4NCI-Frederick Laboratory Animal Sciences Program

This protocol describes the isolation of double-negative thymocytes from the mouse thymus followed by retroviral transduction and co-culture on the delta-like 4-expressing bone marrow stromal cell line co-culture system (OP9-DL4) for further functional analysis.

image

Immunology and Infection

Efficient Transfection of In vitro Transcribed mRNA in Cultured Cells Using Peptide-Poloxamine Nanoparticles
Qin Xiao *1, Yuheng Liu *1, Dandan Zhang 1, Chao Li 1, Qihua Yang 1, Dongshui Lu 1, Weijun Zhang 1, Joseph Rosenecker 2, Quanming Zou 1, Yang Li 3, Shan Guan 1
1National Engineering Research Center of Immunological Products, Department of Microbiology and Biochemical Pharmacy, Third Military Medical University, 2Department of Pediatrics, Ludwig-Maximilians University of Munich, 3Department of Pharmacy, Southwest Hospital, Third Military Medical University

A self-assembled peptide-poloxamine nanoparticle (PP-sNp) is developed using a microfluidic mixing device to encapsulate and deliver in vitro transcribed messenger RNA. The described mRNA/PP-sNp could efficiently transfect cultured cells in vitro.

image

Developmental Biology

Optogenetic Signaling Activation in Zebrafish Embryos
Allison J. Saul *1, Catherine E. Rogers *1, Marcial Garmendia-Cedillos 2, Thomas Pohida 2, Katherine W. Rogers 1
1Division of Developmental Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, NIH, 2Instrumentation Development and Engineering Application Solutions, National Institute of Biomedical Imaging and Bioengineering, NIH

Optogenetic manipulation of signaling pathways can be a powerful strategy to investigate how signaling is decoded in development, regeneration, homeostasis, and disease. This protocol provides practical guidelines for using light-oxygen-voltage sensing domain-based Nodal and bone morphogenic protein (BMP) signaling activators in the early zebrafish embryo.

JoVE Logo

Riservatezza

Condizioni di utilizzo

Politiche

Ricerca

Didattica

CHI SIAMO

Copyright © 2024 MyJoVE Corporation. Tutti i diritti riservati