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University of Southern California

73 ARTICLES PUBLISHED IN JoVE

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Biology

Assaying the Ability of Diffusible Signaling Molecules to Reorient Embryonic Spinal Commissural Axons
Virginia M. Hazen *1,2, Keith Phan *1, Ken Yamauchi *1,2, Samantha J. Butler 1,2
1Department of Biological Sciences, University of Southern California, 2Neuroscience Graduate Program, University of Southern California

This assay assesses the ability of a signaling molecule, here Bone Morphogenetic Protein 7 (BMP7), to reorient commissural axons. An explant of embryonic dorsal spinal cord is cultured adjacent to an aggregate of COS cells secreting the candidate growth factors. Reoriented commissural axons growing within the explant are visualized by immunohistochemistry.

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Neuroscience

Patch Clamp Recordings from Mouse Retinal Neurons in a Dark-adapted Slice Preparation
A. Cyrus Arman 1, Alapakkam P. Sampath 2
1Neurosciences Graduate Program, University of Southern California, 2Zilkha Neurogenetic Institute, Department of Physiology and Biophysics, University of Southern California Keck School of Medicine

Here we describe a procedure for generating dark-adapted slices of the mouse retina for electrophysiological recordings.

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Biology

Crystallization of Membrane Proteins in Lipidic Mesophases
Wei Liu 1, Vadim Cherezov 1
1Molecular Biology, The Scripps Research Institute

The protocols describe the essential steps for obtaining diffraction quality crystals of a membrane protein starting from reconstitution of the protein in a lipidic cubic phase (LCP), finding initial conditions with LCP-FRAP pre-crystallization assays, setting up LCP crystallization trials and harvesting crystals.

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Biology

Drosophila Pupal Abdomen Immunohistochemistry
Wei Wang 1, John H. Yoder 1
1Department of Biological Sciences, University of Alabama

Antibody staining of the Drosophila pupae can enhance genetic analyses of adult abdominal developmental genetics. We present our protocol for dissection, fixation and antibody staining of staged Drosophila pupal abdomen.

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Neuroscience

Cross-Modal Multivariate Pattern Analysis
Kaspar Meyer 1, Jonas T. Kaplan 1
1Brain and Creativity Institute and Department of Psychology, University of Southern California

Classical multivariate pattern analysis predicts sensory stimuli a subject perceives from neural activity in the corresponding cortices (e.g. visual stimuli from activity in visual cortex). Here, we apply pattern analysis cross-modally and show that sound- and touch-implying visual stimuli can be predicted from activity in auditory and somatosensory cortices, respectively.

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JoVE Core

Enabling High Grayscale Resolution Displays and Accurate Response Time Measurements on Conventional Computers
Xiangrui Li 1, Zhong-Lin Lu 2,3,4,5
1Center for Cognitive and Behavioral Brain Imaging, The Ohio State University, 2Department of Psychology, University of Southern California, 3Biomedical Engineering, University of Southern California, 4Neuroscience Graduate Program, University of Southern California, 5Department of Psychology, The Ohio State University

Conventional computer hardware can not generate visual stimuli with sufficiently high grayscale resolution and measure response times with sufficient accuracy. We describe how to use the VideoSwitcher to produce high-resolution monochromatic displays, and the RTbox to measure response times with high accuracy on conventional computer hardware.

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Medicine

Doppler Optical Coherence Tomography of Retinal Circulation
Ou Tan 1, Yimin Wang 1, Ranjith K. Konduru 2, Xinbo Zhang 1, SriniVas R. Sadda 2, David Huang 1
1Department of Ophthalmology, Oregon Health and Science University , 2Department of Ophthalmology, University of Southern California

Total retinal blood flow is measured by Doppler optical coherence tomography and semi-automated grading software.

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Engineering

Fabrication of Silica Ultra High Quality Factor Microresonators
Ashley J. Maker 1, Andrea M. Armani 1,2
1Department of Chemical Engineering and Materials Science, University of Southern California, 2Department of Electrical Engineering-Electrophysics, University of Southern California

We describe the use of a carbon dioxide laser reflow technique to fabricate silica resonant cavities, including free-standing microspheres and on-chip microtoroids. The reflow method removes surface imperfections, allowing long photon lifetimes within both devices. The resulting devices have ultra high quality factors, enabling applications ranging from telecommunications to biodetection.

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Medicine

Imaging Glioma Initiation In Vivo Through a Polished and Reinforced Thin-skull Cranial Window
Lifeng Zhang *1, Andree Lapierre *1, Brittany Roy 1, Maili Lim 1, Jennifer Zhu 1, Wei Wang 1, Stephen B. Sampson 1, Kyuson Yun 1, Bonnie Lyons 1, Yun Li 1, Da-Ting Lin 1
1The Jackson Laboratory

By combining a polished and reinforced thin-skull (PoRTS) cranial window and glioblastoma (GBM) cell injection, we can observe glioma initiation and growth from injected GBM cells in the brain of a live mouse longitudinally.

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Medicine

Heterotypic Three-dimensional In Vitro Modeling of Stromal-Epithelial Interactions During Ovarian Cancer Initiation and Progression
Kate Lawrenson 1, Barbara Grun 2, Simon A. Gayther 1
1Department of Preventive Medicine, University of Southern California, 2Institute for Women's Health, University College London

We describe methodologies for establishing in vitro heterotypic three-dimensional models comprising ovarian fibroblasts and normal ovarian surface or ovarian cancer epithelial cells. We discuss the use of these models to study stromal-epithelial interactions that occur during ovarian cancer development.

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JoVE Journal

Use of LysoTracker to Detect Programmed Cell Death in Embryos and Differentiating Embryonic Stem Cells
Jennifer L. Fogel 1, Thu Zan Tun Thein 1, Francesca V. Mariani 1
1Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, Keck School of Medicine, University of Southern California

We present a simple protocol to visualize regions of programmed cell death (PCD) in mouse embryos and differentiating embryonic stem (ES) cell cultures using a highly soluble dye called LysoTracker.

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Bioengineering

Integrated Photoacoustic Ophthalmoscopy and Spectral-domain Optical Coherence Tomography
Wei Song *1,2, Qing Wei *1, Shuliang Jiao 3, Hao F. Zhang 1,4
1Department of Biomedical Engineering, Northwestern University, 2Department of Physics, Harbin Institute of Technology, 3Department of Ophthalmology, University of Southern California, 4Department of Ophthalmology, Northwestern University

Photoacoustic ophthalmology (PAOM), an optical-absorption-based imaging modality, provides the complementary evaluation of the retina to the currently available ophthalmic imaging technologies. We report the using of PAOM integrated with spectral-domain optical coherence tomography (SD-OCT) for simultaneous multimodal retinal imaging in rats.

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Neuroscience

Imaging pHluorin-tagged Receptor Insertion to the Plasma Membrane in Primary Cultured Mouse Neurons
Yun Li 1, Brittany D. Roy 1, Wei Wang 1, Lifeng Zhang 1, Stephen B. Sampson 1, Da-Ting Lin 1
1The Jackson Laboratory

By tagging the extracellular domains of membrane receptors with superecliptic pHluorin, and by imaging these fusion receptors in cultured mouse neurons, we can directly visualize individual vesicular insertion events of the receptors to the plasma membrane. This technique will be instrumental in elucidating the molecular mechanisms governing receptor insertion to the plasma membrane.

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Bioengineering

Analysis of Targeted Viral Protein Nanoparticles Delivered to HER2+ Tumors
Jae Youn Hwang 1, Daniel L. Farkas 1, Lali K. Medina-Kauwe 2,3
1Department of Biomedical Engineering, University of Southern California, 2Department of Biomedical Sciences, Cedars-Sinai Medical Center, 3Geffen School of Medicine, University of California, Los Angeles

This article details the procedures for optical imaging analysis of the tumor-targeted nanoparticle, HerDox. In particular, detailed use of the multimode imaging device for detecting tumor targeting and assessing tumor penetration is described here.

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Medicine

The Goeckerman Regimen for the Treatment of Moderate to Severe Psoriasis
Rishu Gupta 1,2, Maya Debbaneh 2,3, Daniel Butler 2,4, Monica Huynh 2,5, Ethan Levin 2, Argentina Leon 2, John Koo 2, Wilson Liao 2
1Keck School of Medicine, University of Southern California, 2Psoriasis and Skin Treatment Center Dermatology, University of California, San Francisco , 3University of California Irvine School of Medicine, 4University of Arizona College of Medicine, 5Chicago College of Osteopathic Medicine

Psoriasis is a chronic, immune-mediated inflammatory skin disease. The Goeckerman regimen, formulated for the treatment of psoriasis, consists of exposure to ultraviolet B (UVB) light and application of crude coal tar (CCT). The following protocol is for the administration of Goeckerman therapy for the treatment of moderate-to-severe psoriasis.

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JoVE Journal

A Microfluidic Technique to Probe Cell Deformability
David J. Hoelzle 1,2, Bino A. Varghese 1,3, Clara K. Chan 1, Amy C. Rowat 1
1Department of Integrative Biology and Physiology, University of California, Los Angeles, 2Department of Aerospace and Mechanical Engineering, University of Notre Dame, 3Molecular Imaging Center, University of Southern California

We demonstrate a microfluidics-based assay to measure the timescale for cells to transit through a sequence of micron-scale constrictions.

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Bioengineering

Development of Amelogenin-chitosan Hydrogel for In Vitro Enamel Regrowth with a Dense Interface
Qichao Ruan 1, Janet Moradian-Oldak 1
1Center for Craniofacial Molecular Biology, Herman Ostrow School of Dentistry, University of Southern California

In this article, we describe a protocol for fabricating an amelogenin-chitosan hydrogel for superficial enamel reconstruction. Organized in situ growth of apatite crystals in the hydrogel formed a dense enamel-restoration interface, which will improve the effectiveness and durability of restorations.

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Bioengineering

Rapid and Low-cost Prototyping of Medical Devices Using 3D Printed Molds for Liquid Injection Molding
Philip Chung 1, J. Alex Heller 1, Mozziyar Etemadi 1, Paige E. Ottoson 2, Jonathan A. Liu 3, Larry Rand 2, Shuvo Roy 1
1Department of Bioengineering & Therapeutic Sciences, University of California, San Francisco, 2Department of Obstetrics, Gynecology & Reproductive Sciences, University of California, San Francisco, 3Keck School of Medicine, University of Southern California

We have devised a method for low-cost and rapid prototyping of liquid elastomer rubber injection molded devices by using fused deposition modeling 3D printers for mold design and a modified desiccator as a liquid injection system.

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Medicine

A Surgical Procedure for Resecting the Mouse Rib: A Model for Large-Scale Long Bone Repair
Nikita Tripuraneni *1, Marissa K. Srour *1, John W. Funnell 1, Thu Zan Tun Thein 1, Francesca V. Mariani 1
1Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research, Keck School of Medicine, University of Southern California

The overall goal of this procedure is to successfully resect a portion of bone from the rib of a mouse. The procedure was developed as a model to study large-scale long bone repair.

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Medicine

Three-dimensional Co-culture Model for Tumor-stromal Interaction
Masafumi Horie 1,2, Akira Saito 1,3, Yoko Yamaguchi 4, Mitsuhiro Ohshima 5, Takahide Nagase 1
1Department of Respiratory Medicine, Graduate School of Medicine, The University of Tokyo, 2Department of Clinical Laboratory, Graduate School of Medicine, The University of Tokyo, 3Division for Health Service Promotion, The University of Tokyo, 4Department of Biochemistry, Nihon University School of Dentistry, 5Department of Biochemistry, Ohu University School of Pharmaceutical Sciences

Here we present a protocol to co-culture in three-dimensions, which is useful for investigating multicellular interactions and extracellular matrix-dependent modulation of cancer cell behavior. In this experimental model, cancer cells are cultured on collagen gels embedded with human cancer-associated fibroblasts.

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Medicine

Whole Vitreous Humor Dissection for Vitreodynamic Analysis
Karthik Murali 1, Amir H. Kashani 2, Mark S. Humayun 1
1Biomedical Engineering, University of Southern California, 2Ophthalmology, University of Southern California

The goal of this protocol is to show an effective technique to isolate whole, intact vitreous core and cortex from post mortem enucleated porcine eyes.

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Behavior

Quantifying Learning in Young Infants: Tracking Leg Actions During a Discovery-learning Task
Barbara Sargent 1, Hendrik Reimann 2, Masayoshi Kubo 3, Linda Fetters 1
1Division of Biokinesiology & Physical Therapy at the Herman Ostrow School of Dentistry, University of Southern California, 2Department of Kinesiology, Temple University, 3Department of Physical Therapy, Niigata University of Health and Welfare

A method is described in which 3-4 month old infants learn a task by discovery and their leg movements are captured to quantify the learning process.

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Chemistry

Facile and Efficient Preparation of Tri-component Fluorescent Glycopolymers via RAFT-controlled Polymerization
Wei Wang 1, John M. Lester 1, Anthony E. Amorosa 2, Deborah L. Chance 3, Valeri V. Mossine 1, Thomas P. Mawhinney 1,4
1Department of Biochemistry, University of Missouri, 2Department of Biological Sciences, University of Missouri, 3Department of Molecular Microbiology & Immunology, University of Missouri, 4Department of Child Health, University of Missouri

An efficient, three-step synthesis of RAFT-based fluorescent glycopolymers, consisting of glycomonomer preparation, copolymerization, and post-modification, is demonstrated. This protocol can be used to prepare RAFT-based statistical glycopolymers with desired structures.

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Medicine

Utilizing an Orally Dissolving Strip for Pharmacological and Toxicological Studies: A Simple and Humane Alternative to Oral Gavage for Animals
Nhat Huynh *1, Natalie Arabian *1, Dustin Lieu 1, Liana Asatryan 1, Daryl L. Davies 1
1Titus Family Department of Clinical Pharmacy and Pharmaceutical Economics & Policy, University of Southern California School of Pharmacy

The following protocol describes a novel method for chronic oral drug administration using an orally dissolving strip (ODS) in lieu of the more commonly used oral gavage method. We demonstrate that preclinical, oral drug delivery using the ODS method represents a safe, convenient, and humane alternative to oral gavage.

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Biology

Genome Editing in Astyanax mexicanus Using Transcription Activator-like Effector Nucleases (TALENs)
Johanna E. Kowalko 1, Li Ma 2, William R. Jeffery 3
1Genetics, Development and Cell Biology, Iowa State University, 2Department of Biological Sciences, University of Cincinnati, 3Department of Biology, University of Maryland

Gene-targeting mutagenesis is now possible in a wide range of organisms using genome editing techniques. Here, we demonstrate a protocol for targeted gene mutagenesis using transcription activator like effector nucleases (TALENs) in Astyanax mexicanus, a species of fish that includes surface fish and cavefish.

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Biochemistry

Preparation and Delivery of Protein Microcrystals in Lipidic Cubic Phase for Serial Femtosecond Crystallography
Andrii Ishchenko 1,2, Vadim Cherezov 1,2, Wei Liu 3
1The Bridge Institute, University of Southern California, 2Department of Chemistry, University of Southern California, 3School of Molecular Sciences, Center for Applied Structural Discovery at the Biodesign Institute, Arizona State University

We describe procedures for the preparation and delivery of membrane protein microcrystals in lipidic cubic phase for serial crystallography at X-ray free-electron lasers and synchrotron sources. These protocols can also be applied for incorporation and delivery of soluble protein microcrystals, leading to substantially reduced sample consumption compared to liquid injection.

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Education

Characterizing Electron Transport through Living Biofilms
Matthew Yates 1, Sarah Strycharz-Glaven 1, Joel Golden 1, Jared Roy 1,2, Stanislav Tsoi 3, Jeffrey Erickson 1, Mohamed El-Naggar 4, Scott Calabrese Barton 5, Leonard Tender 1
1Center for Bio/Molecular Science and Engineering, Naval Research Laboratory, 2George Mason University, 3Chemistry Division, Naval Research Laboratory, 4Departments of Physics, Biological Sciences, and Chemistry, University of Southern California, 5Department of Chemical Engineering and Materials Science, Michigan State University

A protocol for measuring electrical conductivity of living microbial biofilms under physiologically relevant conditions is presented.

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Neuroscience

Novel Assay for Cold Nociception in Drosophila Larvae
Heather N. Turner 1,2,5, Christian Landry 3, Michael J. Galko 1,2,4
1Department of Genetics, UT MD Anderson Cancer Center, 2Neuroscience Program, Graduate School of Biomedical Sciences at Houston, 3ProDev Engineering, 4Genes and Development Program, Graduate School of Biomedical Sciences at Houston, 5Section of Neurobiology, University of Southern California

Here we demonstrate a novel assay to study cold nociception in Drosophila larvae. This assay utilizes a custom-built Peltier probe capable of applying a focal noxious cold stimulus and results in quantifiable cold-specific behaviors. This technique will allow further cellular and molecular dissection of cold nociception.

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Biochemistry

A Modified Yeast-one Hybrid System for Heteromeric Protein Complex-DNA Interaction Studies
Prateek Tripathi 1, José L. Pruneda-Paz 2, Steve A. Kay 1
1Department of Neurology, University of Southern California, 2Section of Cell and Developmental Biology, University of California San Diego

The modified yeast one-hybrid assay described here is an extension of the classical yeast one-hybrid (Y1H) assay to study and validate the heteromeric protein complex-DNA interaction in a heterologous system for any functional genomics study.

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Cancer Research

Conditional Knockdown of Gene Expression in Cancer Cell Lines to Study the Recruitment of Monocytes/Macrophages to the Tumor Microenvironment
Marta H. Kubala 1,2, Yves A. DeClerck 1,2,3
1Division of Hematology, Oncology and Blood and Marrow Transplantation, Department of Pediatrics, University of Southern California, 2The Saban Research Institute of Children's Hospital Los Angeles, 3Department of Biochemistry and Molecular Medicine, University of Southern California

This protocol serves as a scheme for setting up a functional Tet-ON system in cancer cell lines and its subsequent use, in particular for studying the role of tumor cell-derived proteins in recruitment of monocytes/macrophages to the tumor microenvironment.

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Bioengineering

Quantifying Microorganisms at Low Concentrations Using Digital Holographic Microscopy (DHM)
Manuel Bedrossian 1, Casey Barr 2, Chris A. Lindensmith 3, Kenneth Nealson 2, Jay L. Nadeau 1
1Department of Medical Engineering, California Institute of Technology, 2Department of Earth Sciences, University of Southern California, 3Jet Propulsion Laboratory, California Institute of Technology

Digital holographic microscopy (DHM) is a volumetric technique that allows imaging samples 50-100X thicker than brightfield microscopy at comparable resolution, with focusing performed post-processing. Here DHM is used for identifying, counting, and tracking microorganisms at very low densities and compared with optical density measurements, plate count, and direct count.

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Neuroscience

Establishing Mouse Models for Zika Virus-induced Neurological Disorders Using Intracerebral Injection Strategies: Embryonic, Neonatal, and Adult
Stephanie A. Herrlinger 1, Qiang Shao 2, Li Ma 2, Melinda Brindley 3, Jian-Fu Chen 2
1Biomedical and Health Sciences Institute, University of Georgia, 2Center for Craniofacial Molecular Biology, University of Southern California, 3Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia

Here we describe a method for establishing a model of Zika virus-induced microcephaly in mouse. This protocol includes methods for embryonic, neonatal, and adult-stage intracerebral inoculation of the Zika virus.

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Biology

Subcellular Fractionation for ERK Activation Upon Mitochondrial-derived Peptide Treatment
Su-Jeong Kim 1, Jialin Xiao 1, Pinchas Cohen 1, Kelvin Yen 1
1Leonard Davis School of Gerontology, University of Southern California

This protocol describes how to stimulate cells with mitochondrial-derived peptides and assess the signaling cascade and localization of phospho-proteins.

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Bioengineering

Synthesis of Monocyte-targeting Peptide Amphiphile Micelles for Imaging of Atherosclerosis
Christopher Poon 1, Manjima Sarkar 1, Eun Ji Chung 1
1Department of Biomedical Engineering, University of Southern California

This paper presents a method involving the synthesis and characterization of monocyte-targeting peptide amphiphile micelles and the corresponding assays to test for biocompatibility and ability of the micelle to bind to monocytes.

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JoVE Core

Murine Drinking Models in the Development of Pharmacotherapies for Alcoholism: Drinking in the Dark and Two-bottle Choice
Nhat Huynh *1, Natalie M Arabian *1, Liana Asatryan 1, Daryl L Davies 1
1Titus Family Department of Clinical Pharmacy and Pharmaceutical Economics & Policy, School of Pharmacy, University of Southern California

Alcohol Use Disorder (AUD) is a major national health problem and the development of more effective treatments is required to offset the needs of this patient population. To this end, the following protocol utilizes two simple rodent drinking models to assess the preclinical efficacy of lead anti-alcohol compounds.

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Biology

Quantification of Lipid Abundance and Evaluation of Lipid Distribution in Caenorhabditis elegans by Nile Red and Oil Red O Staining
Wilber Escorcia 1, Dana L. Ruter 1,2,3, James Nhan 1,2, Sean P. Curran 1,2
1Leonard Davis School of Gerontology, University of Southern California, 2Molecular & Computational Biology Section, University of Southern California, 3Integrative Program for Biological & Genome Sciences, University of North Carolina at Chapel Hill

Nile red staining of fixed Caenorhabditis elegans is a method for quantitative measurement of neutral lipid deposits, while oil red O staining facilitates qualitative assessment of lipid distribution among tissues.

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Immunology and Infection

The Lambda Select cII Mutation Detection System
Ahmad Besaratinia 1, Stella Tommasi 1
1Department of Preventive Medicine, USC Keck School of Medicine, University of Southern California

We describe a detailed protocol for the Lambda Select cII mutation assay in cultured cells of transgenic rodents or the corresponding animals treated with a chemical/physical agent of interest. This approach has been widely used for mutagenicity testing of carcinogens in mammalian cells.

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Biology

Molecular Analysis of Endothelial-mesenchymal Transition Induced by Transforming Growth Factor-β Signaling
Hiroshi I. Suzuki 1, Masafumi Horie 2,3, Hajime Mihira 4, Akira Saito 2
1David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, 2Department of Respiratory Medicine, Graduate School of Medicine, The University of Tokyo, 3Hastings Center for Pulmonary Research, Division of Pulmonary, Critical Care and Sleep Medicine, Department of Medicine, Keck School of Medicine, University of Southern California, 4Department of Molecular Pathology, Graduate School of Medicine, The University of Tokyo

A protocol for in vitro induction of endothelial-mesenchymal transition (EndMT), which is useful for investigating cellular signaling pathways involved in EndMT, is described. In this experimental model, EndMT is induced by treatment with TGF-β in MS-1 endothelial cells.

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Environment

Self-standing Electrochemical Set-up to Enrich Anode-respiring Bacteria On-site
Akihiro Okamoto 1, Annette Rowe 2, Xiao Deng 3, Kenneth H. Nealson 2
1International Center for Materials Nanoarchitectonics, National Institute for Materials Science, 2Department of Earth Sciences, University of Southern California, 3Department of Applied Chemistry, The University of Tokyo

On-site microbial enrichment or in situ cultivation techniques can facilitate the isolation of difficult-to-culture microbial taxa, especially from low-biomass or geochemically extreme environments. Here, we describe an electrochemical set-up without using an external power source to enrich microbial strains that are capable of extracellular electron transport (EET).

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Medicine

Murine Oropharyngeal Aspiration Model of Ventilator-associated and Hospital-acquired Bacterial Pneumonia
Travis B. Nielsen 1, Jun Yan 1, Brian Luna 1, Brad Spellberg 1,2
1Department of Molecular Microbiology and Immunology, University of Southern California, 2Division of Infectious Diseases, Department of Medicine, LAC+USC Medical Center

Infectious pneumonia is among the most common infections in human. An appropriate in vivo model is critical for understanding disease pathogenesis and testing the efficacy of novel therapeutics. With this murine oropharyngeal aspiration pneumonia model, one can examine the pathogenesis and new treatments against these deadly infections.

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Cancer Research

Assessment of Resistance to Tyrosine Kinase Inhibitors by an Interrogation of Signal Transduction Pathways by Antibody Arrays
Katrin Tiemann 1, Carolina Garri 1, Jeffrey Wang 1, Lauren Clarke 1, Kian Kani 1
1Lawrence J. Ellison Institute for Transformative Medicine, Center for Applied Molecular Medicine, University of Southern California

Here, we present a protocol for antibody arrays to identify alterations in signaling pathways in various cellular models. These changes, caused by drugs/hypoxia/ultra-violet light/radiation, or by overexpression/downregulation/knockouts, are important for various disease models and can indicate whether a therapy will be effective or can identify mechanisms of drugs resistance.

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Cancer Research

Phosphopeptide Enrichment Coupled with Label-free Quantitative Mass Spectrometry to Investigate the Phosphoproteome in Prostate Cancer
Larry C. Cheng *1,2, Zhen Li *3, Thomas G. Graeber 4, Nicholas A. Graham 5, Justin M. Drake 1,2,3,6,7
1Graduate Program in Cellular and Molecular Pharmacology, School of Graduate Studies, Rutgers University, The State University of New Jersey, 2Graduate Program in Quantitative Biomedicine, School of Graduate Studies, Rutgers University, The State University of New Jersey, 3Department of Medicine, Division of Medical Oncology, Rutgers Robert Wood Johnson Medical School, 4Crump Institute for Molecular Imaging, Department of Molecular and Medical Pharmacology, Jonsson Comprehensive Cancer Center, UCLA Metabolomics Center, and California NanoSystems Institute, David Geffen School of Medicine, University of California, Los Angeles, 5Mork Family Department of Chemical Engineering and Materials Science, University of Southern California, 6Pharmacology, Rutgers Robert Wood Johnson Medical School, 7Cancer Metabolism and Growth Program, Rutgers Cancer Institute of New Jersey

This protocol describes a procedure to extract and enrich phosphopeptides from prostate cancer cell lines or tissues for an analysis of the phosphoproteome via mass spectrometry-based proteomics.

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Genetics

Culturing and Manipulation of O9-1 Neural Crest Cells
Bao H. Nguyen 1, Mamoru Ishii 2, Robert E. Maxson 2, Jun Wang 1
1Molecular Physiology and Biophysics, Baylor College of Medicine, 2Department of Biochemistry and Molecular Biology, University of Southern California

O9-1 is a multipotent mouse neural crest cell line. Here we describe detailed step-by-step protocols for culturing O9-1 cells, differentiating O9-1 cells into specific cell types, and genetically manipulating O9-1 cells by using siRNA-mediated knockdown or CRISPR-Cas9 genome editing.

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Medicine

A Metadata Extraction Approach for Clinical Case Reports to Enable Advanced Understanding of Biomedical Concepts
John Harry Caufield 1,2, David A. Liem 1,2,3, Anders O. Garlid 1,2, Yijiang Zhou 4, Karol Watson 1,3, Alex A. T. Bui 1,5,6,7, Wei Wang 1,7,8,9, Peipei Ping 1,2,3,7,8
1The NIH BD2K Center of Excellence in Biomedical Computing, University of California, Los Angeles, 2Department of Physiology, University of California, Los Angeles, 3Department of Medicine/Cardiology, University of California, Los Angeles, 4Department of Cardiology, First Affiliated Hospital, Zhejiang University School of Medicine, 5Department of Radiological Sciences, University of California, Los Angeles, 6Department of Bioengineering, University of California, Los Angeles, 7Scalable Analytics Institute (ScAi), University of California, Los Angeles, 8Department of Bioinformatics, University of California, Los Angeles, 9Department of Computer Science, University of California, Los Angeles

We present a protocol and associated metadata template for the extraction of text describing biomedical concepts in clinical case reports. The structured text values produced through this protocol can support deep analysis of thousands of clinical narratives.

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Indium Phosphide Synaptic Device on Silicon to Emulate Synaptic Behavior for Neuromorphic Computing
Debarghya Sarkar *1, Jun Tao *1, Wei Wang 1, Qingfeng Lin 1, Matthew Yeung 1, Chenhao Ren, Rehan Kapadia 1
1Ming Hsieh Department of Electrical Engineering, University of Southern California

The protocol describes the fabrication method of a scalable array of indium phosphide channel field-effect transistors from indium phosphide nanostripes grown directly on an Si/SiO2 substrate with a thin MoOx buffer and the characterization of the field-effect transistors as artificial synaptic devices.

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Medicine

Cloud-Based Phrase Mining and Analysis of User-Defined Phrase-Category Association in Biomedical Publications
Dibakar Sigdel *1,2, Vincent Kyi *1,2, Aiden Zhang *1, Shaun P. Setty 3, David A. Liem 1,2,4, Yu Shi 5, Xuan Wang 5, Jiaming Shen 5, Wei Wang 1,6,7, JiaWei Han 5, Peipei Ping 1,2,4,6
1The NIH BD2K Center of Excellence in Biomedical Computing, University of California, Los Angeles, 2Department of Physiology, University of California, Los Angeles, 3Department of Pediatric and Adult Congenital Heart Surgery, Miller Children's and Women's Hospital and Long Beach Memorial Hospital, 4Department of Medicine/Cardiology, University of California, Los Angeles, 5NIH BD2K Program Centers of Excellence for Big Data Computing -- KnowEng Center, Department of Computer Science, University of Illinois at Urbana-Champaign (UIUC), 6Scalable Analytics Institute (ScAi), University of California, Los Angeles, 7Department of Computer Science, University of California, Los Angeles

We present a protocol and associated programming code as well as metadata samples to support a cloud-based automated identification of phrases-category association representing unique concepts in user selected knowledge domain in biomedical literature. The phrase-category association quantified by this protocol can facilitate in depth analysis in the selected knowledge domain.

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Medicine

Orthotopic Rat Kidney Transplantation: A Novel and Simplified Surgical Approach
Ali R. Ahmadi 1, Le Qi 1, Kenichi Iwasaki 1, Wei Wang 1, Russell N. Wesson 1, Andrew M. Cameron 1, Zhaoli Sun 1
1Department of Surgery, Johns Hopkins University School of Medicine

The purpose of this manuscript and protocol is to explain and demonstrate in detail the surgical procedure of orthotopic kidney transplantation in rats. This method is simplified to achieve the correct perfusion of the donor kidney and shorten the reperfusion time by using the venous and ureteral cuff anastomosis technique.

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Environment

A Uniaxial Compression Experiment with CO2-Bearing Coal Using a Visualized and Constant-Volume Gas-Solid Coupling Test System
Weitao Hou 1, Hanpeng Wang 1, Wei Wang 1, Zhongzhong Liu 1, Qingchuan Li 1
1Research Center of Geotechnical and Structural Engineering, Shandong University

This protocol demonstrates how to prepare a briquette sample and conduct a uniaxial compression experiment with a briquette in different CO2 pressures using a visualized and constant-volume gas-solid coupling test system. It also aims to investigate changes in terms of coal’s physical and mechanical properties induced by CO2 adsorption.

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Biology

Examination of Mitotic and Meiotic Fission Yeast Nuclear Dynamics by Fluorescence Live-cell Microscopy
Wilber Escorcia 1,2, Kuo-Fang Shen 1, Ji-Ping Yuan 1, Susan L. Forsburg 1
1Program in Molecular and Computational Biology, University of Southern California, 2Leonard Davis School of Gerontology, University of Southern California

Here, we present live-cell imaging which is a non-toxic microscopy method that allows researchers to study protein behavior and nuclear dynamics in living fission yeast cells during mitosis and meiosis.

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Medicine

Immunoglobulin G N-Glycan Analysis by Ultra-Performance Liquid Chromatography
Di Liu *1, Xizhu Xu *2, Yuejin Li 2, Jie Zhang 1, Xiaoyu Zhang 1, Qihuan Li 1, Haifeng Hou 2, Dong Li 2, Wei Wang 1,2,3, Youxin Wang 1
1Beijing Key Laboratory of Clinical Epidemiology, School of Public Health, Capital Medical University, 2School of Public Health, Shandong First Medical University & Shandong Academy of Medical Sciences, 3School of Medical and Health Sciences, Edith Cowan University

Immunoglobulin G (IgG) N-glycan is characterized using hydrophilic interaction chromatography UPLC. In addition, the structure of IgG N-glycan is clearly separated. Presented here is an introduction to this experimental method so that it can be widely used in research settings.

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Cancer Research

Analyzing Tumor Gene Expression Factors with the CorExplorer Web Portal
Shirley Pepke 1, William M. Nelson 2, Greg Ver Steeg 3
1Lyrid LLC, 2Independent consultant, 3Information Sciences Institute, University of Southern California

We introduce the CorExplorer web portal, an resource for exploration of tumor RNA sequencing factors found by the machine learning algorithm CorEx (Correlation Explanation), and show how factors can be analyzed relative to survival, database annotations, protein-protein interactions, and one another to gain insight into tumor biology and therapeutic interventions.

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Neuroscience

Retinal Vascular Reactivity as Assessed by Optical Coherence Tomography Angiography
Sam Kushner-Lenhoff 1, Bright S. Ashimatey 1, Amir H. Kashani 1,2
1Department of Ophthalmology, USC Roski Eye Institute, 2USC Ginsberg Institute for Biomedical Therapeutics, Keck School of Medicine of the University of Southern California

This article describes a method for measuring retinal vasculature reactivity in vivo with human subjects using a gas breathing provocation technique to deliver vasoactive stimuli while acquiring retinal images.

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Biology

Measurements of Physiological Stress Responses in C. Elegans
Raz Bar-Ziv *1, Ashley E. Frakes *1, Ryo Higuchi-Sanabria *1, Theodore Bolas 1, Phillip A. Frankino 1, Holly K. Gildea 1, Melissa G. Metcalf 1, Andrew Dillin 1
1Department of Molecular and Cell Biology, University of California, Berkeley

Here, we characterize cellular proteotoxic stress responses in the nematode C. elegans by measuring the activation of fluorescent transcriptional reporters and assaying sensitivity to physiological stress.

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Behavior

A Chronic Sleep Fragmentation Model using Vibrating Orbital Rotor to Induce Cognitive Deficit and Anxiety-Like Behavior in Young Wild-Type Mice
Yi Xie 1, Sai-Yue Deng 1, Si-Miao Chen 1, Xue-Jiao Chen 1, Wen-Wen Lai 1, Li-Fang Huang 1, Li Ba 1, Wei Wang 1,2, Feng-Fei Ding 1,3
1Department of Neurology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430000, China, 2Key Laboratory of Neurological Diseases of Chinese Ministry of Education, The School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430000,China, 3Department of Pharmacology, School of Basic Medical Science; State Key Laboratory of Medical Neurobiology, Institutes of Brain Science and Collaborative Innovation Center for Brain Science, Fudan University, Shanghai 200032, China

Presented here is a protocol for chronic sleep fragmentation (CSF) model achieved by an electrically controlled orbital rotor, which could induce confirmed cognitive deficit and anxiety-like behavior in young wild-type mice. This model can be applied to explore the pathogenesis of chronic sleep disturbance and related disorders.

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Developmental Biology

Incremental Temperature Changes for Maximal Breeding and Spawning in Astyanax mexicanus
Li Ma 1, Ruby Dessiatoun 1, Janet Shi 1, William R. Jeffery 1
1Department of Biology, University of Maryland

This article outlines the basic laboratory conditions and protocols for an incremental temperature regime to stimulate maximal spawning in the Mexican tetra Astyanax mexicanus, which is an emerging model for developmental and evolutionary studies.

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Biology

Xenopus laevis Egg Extract Preparation and Live Imaging Methods for Visualizing Dynamic Cytoplasmic Organization
Xianrui Cheng 1,2, James E. Ferrell, Jr. 1,3
1Department of Chemical and Systems Biology, Stanford University School of Medicine, 2Current Address: Department of Biological Sciences, University of Southern California, 3Department of Biochemistry, Stanford University School of Medicine

We describe a method for the preparation and live imaging of undiluted cytoplasmic extracts from Xenopus laevis eggs.

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Neuroscience

Establishment of a Rat Model of Superior Sagittal-Sinus Occlusion via a Thread-Embolism Method
Weiyan Jiang *1, Congcong Jin *2, Weiming Xu *3, Yingxian Li 4, Yinghong Lin 5, Shengxiang Liang 6,7, Wei Wang 8
1Department of Clinical Laboratory, the Second Affiliated Hospital and Yuying Children’s Hospital of Wenzhou Medical University, 2Rehabilitation Department, Wenzhou Hospital of Integrated Traditional Chinese and Western Medicine, 3Department of Neurosurgery, the Hospital of Changle, 4Pediatrics Ward 2, Shouguang Hospital of Traditional Chinese Medicine, 5Fujian University of Traditional Chinese Medicine, 6National-Local Joint Engineering Research Center of Rehabilitation Medicine Technology, Fujian University of Traditional Chinese Medicine, 7Rehabilitation Industry Institute, Fujian University of Traditional Chinese Medicine, 8Department of Neurosurgery, the First Affiliated Hospital of Wenzhou Medical University

Here, we establish a novel Sprague-Dawley (SD) rat model of superior sagittal sinus (SSS) thrombosis via a thread-embolization method, and the stability and reliability of the model were verified.

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Cancer Research

The Establishment and Utilization of Patient Derived Xenograft Models of Central Nervous System Metastasis
Ben Yi Tew 1, Bodour Salhia 1
1Department of Translational Genomics, Keck School of Medicine, University of Southern California

Central nervous system metastasis PDX models represent the phenotypic and molecular characteristics of human metastasis, making them excellent models for preclinical studies. Described here is how to establish PDX models and the inoculation routes that are best used for preclinical studies.

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JoVE Journal

Construction of Model Lipid Membranes Incorporating G-protein Coupled Receptors (GPCRs)
Ahmed Elbaradei *1, Lucia Caterina Dalle Ore *1, Noah Malmstadt 1
1Mork Family Department of Chemical Engineering and Materials Science, University of Southern California

This protocol utilizes agarose swelling as a powerful and generalizable technique for incorporating integral membrane proteins (IMPs) into giant unilamellar lipid vesicles (GUVs), as described here for the reconstitution of the human 1A serotonin receptor protein (5-HT1AR), one of the classes of pharmacologically important G protein-coupled receptors.

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Medicine

Robot-assisted Total Mesorectal Excision and Lateral Pelvic Lymph Node Dissection for Locally Advanced Middle-low Rectal Cancer
Chenhao Hu *1,2,3, Zhe Zhang *1,2,3, Lei Zhang 1,2,3, Ruihan Liu 2,3, Jun Yan 1,2,3, Qi Sun 1,2,3, Guanghui Wang 1, Junjun She 1,2,3
1Department of General Surgery, The First Affiliated Hospital of Xian Jiaotong University, 2Center for Gut Microbiome Research, Med-X Institute, The First Affiliated Hospital of Xian Jiao tong University, 3Department of High Talent, The First Affiliated Hospital of Xian Jiaotong University

The robotic technique described herein aims to detail a stepwise approach to robot-assisted total mesorectal excision and lateral pelvic lymph node dissection for locally advanced (T3/T4) rectal cancer located below the peritoneal reflection.

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Cancer Research

Aqueous Humor as a Liquid Biopsy for Retinoblastoma: Clear Corneal Paracentesis and Genomic Analysis
Mary E. Kim 1,2, Liya Xu 1,2,3,11, Rishvanth K. Prabakar 4, Lishuang Shen 5, Chen-Ching Peng 1,3, Peter Kuhn 3,6,7,8, Xiaowu Gai 5,9, James Hicks 3,6,10, Jesse L. Berry 1,2,6,11
1The Vision Center, Children’s Hospital Los Angeles, 2USC Roski Eye Institute, Keck School of Medicine of the University of Southern California, 3Department of Biological Sciences, Dornsife College of Letters, Arts, and Sciences, University of Southern California, 4Department of Molecular and Computational Biology, University of Southern California, 5Center for Personalized Medicine, Department of Pathology and Laboratory Medicine, Children’s Hospital Los Angeles, 6Norris Comprehensive Cancer Center, Keck School of Medicine, University of Southern California, 7Department of Aerospace and Mechanical Engineering, Viterbi School of Engineering, University of Southern California, 8Department of Biomedical Engineering, Viterbi School of Engineering, University of Southern California, 9Department of Pathology and Laboratory Medicine, Keck School of Medicine of USC, 10Department of Biochemistry and Molecular Medicine, Keck School of Medicine, University of Southern California, 11The Saban Research Institute, Children’s Hospital Los Angeles

The aqueous humor is a high-yield liquid biopsy for retinoblastoma, intraocular cancer that cannot be biopsied in vivo due to the risk of extraocular spread. Herein, a method for safely extracting aqueous humor via clear corneal paracentesis and steps for genomic analysis to identify prognostic biomarkers are presented.

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Neuroscience

Two Peeling Methods for the Isolation of Photoreceptor Cell Compartments in the Mouse Retina for Protein Analysis
Kasey Rose 1, Sowmya Lokappa 1, Jeannie Chen 1,2
1Zilkha Neurogenetic Institute, Keck School of Medicine, University of Southern California, 2Department of Cell & Neurobiology, Keck School of Medicine, University of Southern California

This protocol presents two techniques to isolate the subcellular compartments of murine rod photoreceptors for protein analysis. The first method utilizes live retinae and cellulose filter paper to separate rod outer segments, while the second employs lyophilized retinae and adhesive tape to peel away rod inner and outer segment layers.

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Neuroscience

A New Optokinetic Testing Method to Measure Rat Vision
Faizah Ahmed *1, Deepthi S. Rajendran Nair *1, Biju B. Thomas 1,2
1Roski Eye Institute, Department of Ophthalmology, Keck School of Medicine, University of Southern California, 2USC Ginsburg Institute for Biomedical Therapeutics, University of Southern California

The Optokinetic Nystagmus (OKN) behavioral testing method is used for the assessment of visual acuity in rodents. Here a simple method is demonstrated that can be easily set up in research laboratories for a reliable assessment of visual function in both normal and experimental rats.

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Genetics

Surveying Low-Cost Methods to Measure Lifespan and Healthspan in Caenorhabditis elegans
Toni Castro Torres *1, Darius Moaddeli *1, Maxim Averbukh 1, Aeowynn J. Coakley 1, Naibedya Dutta 1, Gilberto Garcia 1, Ryo Higuchi-Sanabria 1
1Leonard Davis School of Gerontology, University of Southern California

Caenorhabditis elegans serve as an excellent model system with robust and low-cost methods for surveying healthspan, lifespan, and resilience to stress.

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Neuroscience

Gentle Isolation of Nuclei from the Brain Tissue of Adult African Turquoise Killifish, a Naturally Short-Lived Model for Aging Research
Bryan B. Teefy 1, Ari Adler 1, Rajyk Bhala 1, Bérénice A. Benayoun 1,2,3,4,5
1Leonard Davis School of Gerontology, University of Southern California, 2Molecular and Computational Biology Department, USC Dornsife College of Letters, Arts and Sciences, 3Biochemistry and Molecular Medicine Department, USC Keck School of Medicine, 4Epigenetics and Gene Regulation, USC Norris Comprehensive Cancer Center, 5USC Stem Cell Initiative

Here we present a protocol to isolate nuclei from the brains of the short-lived vertebrate model Nothobranchius furzeri for downstream applications such as single-nucleus RNA sequencing or single-nucleus assay for transposase-accessible chromatin with sequencing (ATAC-seq).

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Neuroscience

Social Isolation Model: A Noninvasive Rodent Model of Stress and Anxiety
Saki Watanabe 1, Alzahra Al Omran 1, Amy S. Shao 2, Jing Liang 1
1Titus Family Department of Clinical Pharmacy, School of Pharmacy, University of Southern California, 2Homer Stryker M.D. School of Medicine, Western Michigan University

Presented here is a social isolation (SI)-induced anxiety mouse model that utilizes wild type C56BL/6J mice to induce stress and anxiety-like behavior with minimal handling and no invasive procedures. This model reflects modern life patterns of social isolation and is ideal for studying anxiety and related disorders.

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Medicine

Development of a Surgical Technique for Subretinal Implants in Rats
Juan Carlos Martinez Camarillo 1,2, Yuntao Hu 1,5, Biju B. Thomas 1,2, Danhong Zhu 3, David R. Hinton 2,3, Debbie Mitra 1,2, Jose Miguel Mora Correa 2, Deepthi S. Rajendran Nair 1,2, Jane Lebkowski 4, Mark S. Humayun 1,2
1USC Roski Eye Institute, Keck School of Medicine of the University of Southern California, 2USC Dr. Allen and Charlotte Ginsburg Institute for Biomedical Therapeutics, University of Southern California, 3Department of Pathology, Keck School of Medicine of the University of Southern California, 4Regenerative Patch Technologies LLC, 5Eye center, Beijing Tsinghua Changgung Hospital, School of Clinical Medicine, Tsinghua University

The present protocol describes the scleral approach for subretinal device implantation, a feasible surgical technique for implementation in animal models of retinal diseases in research.

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Cancer Research

Generating and Imaging Mouse and Human Epithelial Organoids from Normal and Tumor Mammary Tissue Without Passaging
Serena L. Cornelius 1,2, Megan M. Colonnetta 2,3,4,5, Katherine E. Lake 1,2, Clayton A. Smith 1,2, Yu-An Zhang 1,2, Evanthia T. Roussos-Torres 6, Sangeetha M. Reddy 1,2, Elizabeth H. Chen 2,3,4,5, Isaac S. Chan 1,2,3,4
1Department of Internal Medicine, Division of Hematology and Oncology, University of Texas Southwestern Medical Center, 2Harold C. Simmons Comprehensive Cancer Center, University of Texas Southwestern Medical Center, 3Department of Molecular Biology, University of Texas Southwestern Medical Center, 4Hamon Center for Regenerative Science and Medicine, University of Texas Southwestern Medical Center, 5Department of Cell Biology, University of Texas Southwestern Medical Center, 6Division of Medical Oncology, Norris Comprehensive Cancer Center, Keck School of Medicine, University of Southern California

This protocol discusses an approach for generating epithelial organoids from primary normal and tumor mammary tissue through differential centrifugation. Furthermore, instructions are included for three-dimensional culturing as well as immunofluorescent imaging of embedded organoids.

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Neuroscience

Isolating and Culturing Vestibular and Spiral Ganglion Somata from Neonatal Rodents for Patch-Clamp Recordings
Megana R. Iyer 1, Christopher Ventura 1, Daniel Bronson 1, Nathaniel Nowak 1, Katherine Regalado 1, Radha Kalluri 1
1Caruso Department of Otolaryngology Head & Neck Surgery, Zilkha Neurogenetics Institute, Hearing and Communications Neuroscience Training Program, University of Southern California

Presented here are methods providing detailed instructions for dissecting, dissociating, culturing, and patch-clamp recording from vestibular ganglion and spiral ganglion neurons of the inner ear.

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Developmental Biology

Using Avian Skin Explants to Study Tissue Patterning and Organogenesis
Tingxin Jiang 1, Maeve Secor 2, Rusty Lansford 3, Randall B. Widelitz 1, Cheng Ming Chuong 1
1Department of Pathology, Keck School of Medicine of University of Southern California, 2Molecular and Computational Biology, University of Southern California, 3Department of Radiology, Keck School of Medicine of University of Southern California

Here we describe protocols for three types of avian embryonic skin explant cultures that can be used to examine tissue interactions, 4D imaging timelapse movie (3D plus time), global or local perturbation of molecular function, and systems biology characterization.

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Biology

Obtaining High-Quality Cryosections of Whole Rabbit Eye
Erik A. Souverein 1, Aaron Nagiel 2,3,4, Ksenia Gnedeva 5,6
1Keck School of Medicine, University of Southern California, 2The Vision Center, Department of Surgery, Children’s Hospital Los Angeles, 3The Saban Research Institute, Children’s Hospital Los Angeles, 4USC Roski Eye Institute, Department of Ophthalmology, Keck School of Medicine, University of Southern California, 5Eli and Edythe Broad CIRM Center for Regenerative Medicine and Stem Cell Research, University of Southern California, 6Tina and Rick Caruso Department of Otolaryngology-Head and Neck Surgery, University of Southern California

This protocol describes a reliable method for obtaining high-quality cryosections of whole rabbit eyes. It details rabbit eye dissection, fixation, embedding, and sectioning procedures, which may be easily adapted for use in any study utilizing immunohistochemistry in larger eyes.

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Medicine

Determination of Continuity Index Values in Atrial Fibrillation Ablation with Proactive Esophageal Cooling
Catherine Lazarus 1, Jacob Sherman 2, Natalie Putzel 3, Tiffany Sharkoski 4, William Zagrodzky 4, Erik Kulstad 4,5, Alex Ro 6, Jose Nazari 6, Westby Fisher 6, Mark Metzl 6
1Northwestern University, 2Washington University in St. Louis, 3University of Southern California, 4Attune Medical, 5University of Texas Southwestern Medical Center, 6NorthShore University Hospital

This protocol describes the methods used to determine the continuity index in patients undergoing pulmonary vein isolation procedures using radiofrequency ablation and demonstrates the differences in continuity index between ablation procedures using proactive esophageal cooling as compared to procedures using traditional luminal esophageal temperature monitoring.

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Medicine

Consistent Delivery of Adeno-Associated Virus via Lateral Tail-Vein Injection in Adult Mice
Fady Guirguis 1,2, Véronique Bolduc 1, Matthew J. Slarve 3, Haiyan Zhou 4,5, Francesco Muntoni 2,4, Carsten G. Bönnemann 1
1Neuromuscular and Neurogenetic Disorders of Childhood Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, 2The Dubowitz Neuromuscular Centre, Molecular Neurosciences Section, Developmental Neurosciences Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London, 3Department of Molecular Microbiology and Immunology, University of Southern California, 4NIHR Great Ormond Street Hospital Biomedical Research Centre, 5Genetics and Genomic Medicine Research and Teaching Department, Great Ormond Street Institute of Child Health, University College London

Here we detail an optimized protocol for mouse lateral tail-vein injection to systemically administer adeno-associated virus (AAV) in adult mice. Additionally, we describe protocols of commonly used assays to assess AAV transduction.

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