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70 ARTICLES PUBLISHED IN JoVE

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Biology

Isolation and Enrichment of Rat Mesenchymal Stem Cells (MSCs) and Separation of Single-colony Derived MSCs
Linxia Zhang 1, Christina Chan 1
1Department of Chemical Engineering and Materials Science, City of Hope Cancer Center

Rat MSCs were isolated from femurs and tibias and then enriched by magnetic cell sorting. Sorted cells were confirmed for the expression of surface markers by flow cytometry. These cells were also cultured at clonal density to form single colonies and then these colonies were separated by cloning cylinders.

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Biology

Arabidopsis thaliana Polar Glycerolipid Profiling by Thin Layer Chromatography (TLC) Coupled with Gas-Liquid Chromatography (GLC)
Zhen Wang 1, Christoph Benning 1
1Department of Biochemistry and Molecular Biology, Michigan State University

Composition of polar lipid extracts and the fatty acid composition of individual glycerolipids are determined in a simple and robust lipid profiling experiment. For this purpose, glycerolipids are isolated by thin layer chromatography and subjected to transmethylation of their acyl groups. Fatty acyl methylesters are quantified by gas-liquid chromatography.

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Biology

Experimental Manipulation of Body Size to Estimate Morphological Scaling Relationships in Drosophila
R. Craig Stillwell 1, Ian Dworkin 2, Alexander W. Shingleton 2, W. Anthony Frankino 1
1Department of Biology & Biochemistry, University of Houston, 2Department of Zoology, Michigan State University

Morphological scaling relationships capture and describe organismal shape. We present a method to measure morphological scaling relationships across the natural range of body sizes in fully metamorphic insects. Using a simple diet manipulation we increase the distribution of trait sizes, permitting the accurate description of how shape and size co-vary.

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Bioengineering

GENPLAT: an Automated Platform for Biomass Enzyme Discovery and Cocktail Optimization
Jonathan Walton 1,2, Goutami Banerjee 2, Suzana Car 2
1DOE Plant Research Laboratory, Michigan State University, 2DOE Great Lakes Bioenergy Research Center, Michigan State University

GENPLAT (GLBRC Enzyme Platform) is an automated platform for discovery and optimization of enzyme cocktails for biomass degradation. It can be adapted to multiple feedstocks and mixtures of enzymes containing multiple components.

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Medicine

Femoral Arterial and Venous Catheterization for Blood Sampling, Drug Administration and Conscious Blood Pressure and Heart Rate Measurements
Brian Jespersen 1, Lauren Knupp 1, Carrie A. Northcott 1
1Department of Pharmacology and Toxicology, Michigan State University

Chronic catheterization of blood vessels in the rat is often required for administration of substances, obtain blood sample over a period of time or for direct conscious blood pressure measurements. Femoral arterial catheterization of the rat and corresponding measurements of blood pressure in the conscious animal will be demonstrated.

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Biology

Fluorescence-microscopy Screening and Next-generation Sequencing: Useful Tools for the Identification of Genes Involved in Organelle Integrity
Giovanni Stefano 1, Luciana Renna 1, Federica Brandizzi 1
1DOE Plant Research Laboratory, Michigan State University

A fundamental quest in cell biology is to define the mechanisms that underlie the identity of the organelles that make eukaryotic cells. Here we propose a method to identify the genes responsible for the morphological and functional integrity of plant organelles using fluorescence microscopy and next-generation sequencing tools.

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Biology

Sexual Development and Ascospore Discharge in Fusarium graminearum
Brad Cavinder 1, Usha Sikhakolli 2, Kayla M. Fellows 3, Frances Trail 2,4
1Genetics Program, Michigan State University, 2Department of Plant Biology, Michigan State University, 3Human Biology Program, Michigan State University, 4Department of Plant Pathology, Michigan State University

Sexual crosses and isolation of recombinant progeny are important research tools for the filamentous fungus, Fusarium graminearum, The techniques necessary successfully carry out these processes are presented.

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Bioengineering

Microfluidic Mixers for Studying Protein Folding
Steven A. Waldauer 1, Ling Wu 1, Shuhuai Yao 2, Olgica Bakajin 3, Lisa J. Lapidus 1
1Department of Physics and Astronomy, Michigan State University, 2Department of Mechanical Engineering, Hong Kong University of Science and Technology, 3Center for Biophotonics, University of California, Davis

In this work we explain the fabrication and use of a microfluidic mixer capable of mixing two solutions in ~8 μs. We also demonstrate the use of these mixers with spectroscopic detection using UV fluorescence and fluorescence resonance energy transfer (FRET).

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Advanced Biology

Growth Curves: Generating Growth Curves Using Colony Forming Units and Optical Density Measurements
Andrew J. Van Alst 1, Rhiannon M. LeVeque 1, Natalia Martin 1, Victor J. DiRita 1
1Department of Microbiology and Molecular Genetics, Michigan State University

Growth Curves: Generating Growth Curves Using Colony Forming Units and Optical Density Measurements

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Advanced Biology

Microscopy and Staining: Gram, Capsule, and Endospore Staining
Rhiannon M. LeVeque 1, Natalia Martin 1, Andrew J. Van Alst 1, Victor J. DiRita 1
1Department of Microbiology and Molecular Genetics, Michigan State University

Microscopy and Staining: Gram, Capsule, and Endospore Staining

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Advanced Biology

Transformation of E. coli Cells Using an Adapted Calcium Chloride Procedure
Natalia Martin 1, Andrew J. Van Alst 1, Rhiannon M. LeVeque 1, Victor J. DiRita 1
1Department of Microbiology and Molecular Genetics, Michigan State University

Transformation of E. coli Cells Using an Adapted Calcium Chloride Procedure

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JoVE Core

Scanning-probe Single-electron Capacitance Spectroscopy
Kathleen A. Walsh 1, Megan E. Romanowich 1, Morewell Gasseller 1,2, Irma Kuljanishvili 1,3, Raymond Ashoori 4, Stuart Tessmer 1
1Department of Physics and Astronomy, Michigan State University, 2Department of Chemistry & Biochemistry/Physics, Mercyhurst University, 3Department of Physics, Saint Louis University, 4Department of Physics, Massachusetts Institute of Technology

Scanning-probe single-electron capacitance spectroscopy facilitates the study of single-electron motion in localized subsurface regions. A sensitive charge-detection circuit is incorporated into a cryogenic scanning probe microscope to investigate small systems of dopant atoms beneath the surface of semiconductor samples.

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Behavior

A Fully Automated Rodent Conditioning Protocol for Sensorimotor Integration and Cognitive Control Experiments
Ali Mohebi 1, Karim G. Oweiss 1,2,3
1Electrical and Computer Engineering, Michigan State University, 2Neuroscience Program, Michigan State University, 3Cognitive Science Program, Michigan State University

A fully automated protocol for rodent operant conditioning is proposed. The protocol relies on precise temporal control of behavioral events to investigate the extent to which this control influences neural activity underlying sensorimotor integration and cognitive control experiments.

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Bioengineering

Lignin Down-regulation of Zea mays via dsRNAi and Klason Lignin Analysis
Sang-Hyuck Park 1, Rebecca Garlock Ong 2, Chuansheng Mei 3, Mariam Sticklen 4
1The School of Plant Sciences, University of Arizona, 2Department of Chemical Engineering and Materials Science, DOE Great Lakes Bioenergy Research Center, Michigan State University, 3The Institute for Sustainable and Renewable Resources, The Institute for Advanced Learning and Research, 4Department of Plant, Soil and Microbial Sciences, Michigan State University

A double stranded RNA interference (dsRNAi) technique is employed to down-regulate the maize cinnamoyl coenzyme A reductase (ZmCCR1) gene to lower plant lignin content. Lignin down-regulation from the cell wall is visualized by microscopic analyses and quantified by the Klason method. Compositional changes in hemicellulose and crystalline cellulose are analyzed.

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Biology

In Vitro Aggregation Assays Using Hyperphosphorylated Tau Protein
Dexin Sui *1, Mengyu Liu *1, Min-Hao Kuo 1
1Department of Biochemistry and Molecular Biology, Michigan State University

Unmodified and hyperphosphorylated tau proteins were used in two in vitro aggregation assays to reveal the hyperphosphorylation-dependent fast aggregation kinetics. These assays pave the way for future screens for compounds that can modulate the propensity of hyperphosphorylated tau to form fibrils that underlie the progression of Alzheimer’s disease.

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Biology

A New Clarification Method to Visualize Biliary Degeneration During Liver Metamorphosis in Sea Lamprey (Petromyzon marinus)
Yu-Wen Chung-Davidson 1, Peter J. Davidson 1, Anne M. Scott 1, Erin J. Walaszczyk 1, Cory O. Brant 1, Tyler Buchinger 1, Nicholas S. Johnson 2, Weiming Li 1
1Department of Fisheries & Wildlife, Michigan State University, 2Hammond Bay Biological Station, Great Lakes Science Center, U.S. Geological Survey

Sea lamprey lose the gall bladder and bile ducts during metamorphosis, a process similar to human biliary atresia. A new fixation and clarification method (CLARITY) was modified to visualize the entire biliary tree using laser scanning confocal microscopy. This method provides a powerful tool to study biliary degeneration.

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JoVE Core

Measurement of Smooth Muscle Function in the Isolated Tissue Bath-applications to Pharmacology Research
Brian Jespersen 1, Nathan R. Tykocki 2, Stephanie W. Watts 1, Peter J. Cobbett 1
1Department of Pharmacology and Toxicology, Michigan State University, 2Department of Pharmacology, University of Vermont College of Medicine

This protocol describes the measurement of isometric contraction in an isolated smooth muscle preparation, using an isolated tissue bath system and computer-based data acquisition.

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Medicine

Quantitative Magnetic Resonance Imaging of Skeletal Muscle Disease
Bruce M. Damon 1,2,3,4, Ke Li 1,2, Richard D. Dortch 1,2, E. Brian Welch 1,2, Jane H. Park 1,2,4, Amanda K. W. Buck 1,2, Theodore F. Towse 1,2,5, Mark D. Does 1,2,3, Daniel F. Gochberg 1,2,6, Nathan D. Bryant 1,2
1Institute of Imaging Science, Vanderbilt University, 2Department of Radiology and Radiological Sciences, Vanderbilt University, 3Department of Biomedical Engineering, Vanderbilt University, 4Department of Molecular Physiology and Biophysics, Vanderbilt University, 5Department of Physical Medicine and Rehabilitation, Vanderbilt University, 6Department of Physics and Astronomy, Vanderbilt University

Neuromuscular diseases often exhibit a temporally varying, spatially heterogeneous, and multi-faceted pathology. The goal of this protocol is to characterize this pathology using non-invasive magnetic resonance imaging methods.

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JoVE Journal

Physiology Lab Demonstration: Glomerular Filtration Rate in a Rat
Carmen Hinojosa-Laborde 1, Brian Jespersen 2, Robert Shade 3
1Tactical Combat Casualty Care Research, U.S. Army Institute of Surgical Research, 2Department of Pharmacology and Toxicology, Michigan State University, 3Southwest National Primate Research Center, Texas Biomedical Research Institute

The purpose of this protocol is to demonstrate the principles and techniques for measuring and calculating glomerular filtration rate, urine flow rate, and excretion of sodium and potassium in a rat. This demonstration can be used to provide students with an overall conceptual understanding of how to measure renal function.

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Neuroscience

In Situ Ca2+ Imaging of the Enteric Nervous System
David E. Fried 1, Brian D. Gulbransen 1
1Department of Physiology, Michigan State University

The enteric nervous system (ENS) is a network of neurons and glia located in the gut wall that controls intestinal reflexes. This protocol describes methods for recording the activity of enteric neurons and glia in live preparations of ENS using Ca2+ imaging.

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Neuroscience

Functional and Morphological Assessment of Diaphragm Innervation by Phrenic Motor Neurons
Melanie Martin 1,2, Ke Li 1, Megan C. Wright 2, Angelo C. Lepore 1
1Department of Neuroscience, Farber Institute for Neurosciences, Sidney Kimmel Medical College at Thomas Jefferson University, 2Department of Biology, Arcadia University

Compound muscle action potential recording quantitatively assesses functional diaphragm innervation by phrenic motor neurons. Whole-mount diaphragm immunohistochemistry assesses morphological innervation at individual neuromuscular junctions. The goal of this protocol is to demonstrate how these two powerful methodologies can be used in various rodent models of spinal cord disease.

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Bioengineering

Live Cell Imaging during Mechanical Stretch
Gabriel Rápalo 1,2, Josh D. Herwig 3, Robert Hewitt 4, Kristina R. Wilhelm 1,2, Christopher M. Waters 1,2, Esra Roan 3
1Department of Physiology, University of Tennessee Health Science Center, 2Department of Biomedical Engineering and Imaging, University of Tennessee Health Science Center, 3Department of Biomedical Engineering, University of Memphis, 4Department of Engineering Technology, University of Memphis

A novel imaging protocol was developed using a custom motor-driven mechanical actuator to allow the measurement of real time responses to mechanical strain in live cells. Relevant to mechanobiology, the system can apply strains up to 20% while allowing near real-time imaging with confocal or atomic force microscopy.

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Immunology and Infection

Isolation of Leukocytes from the Human Maternal-fetal Interface
Yi Xu 1, Olesya Plazyo 1, Roberto Romero 1,2,3,4, Sonia S. Hassan 1,5, Nardhy Gomez-Lopez 1,5,6
1Perinatology Research Branch, NICHD/NIH/DHHS, 2Department of Obstetrics and Gynecology, University of Michigan, 3Department of Epidemiology and Biostatistics, Michigan State University, 4Department of Molecular Obstetrics and Genetics, Wayne State University, 5Department of Obstetrics and Gynecology, Wayne State University School of Medicine, 6Department of Immunology and Microbiology, Wayne State University School of Medicine

Described herein is a protocol to isolate and further study the infiltrating leukocytes of the decidua basalis and decidua parietalis - the human maternal-fetal interface. This protocol maintains the integrity of cell surface markers and yields enough viable cells for downstream applications as proven by flow cytometry analysis.

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Neuroscience

An Approach to Enhance Alignment and Myelination of Dorsal Root Ganglion Neurons
Chun Liu 1, Christina Chan 1,2
1Department of Chemical Engineering and Materials Science, Michigan State University, 2Department of Biochemistry and Molecular Biology, Michigan State University

This protocol describes the isolation of dorsal root ganglion (DRG) neurons isolated from rats and the culture of DRG neurons on a static pre-stretched cell culture system to enhance axon alignment, with subsequent co-culture of Schwann Cells (SCs) to promote myelination.

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JoVE Journal

Techniques for the Evolution of Robust Pentose-fermenting Yeast for Bioconversion of Lignocellulose to Ethanol
Patricia J. Slininger 1, Maureen A. Shea-Andersh 1, Stephanie R. Thompson 1, Bruce S. Dien 1, Cletus P. Kurtzman 2, Leonardo Da Costa Sousa 3, Venkatesh Balan 3
1Bioenergy Research Unit, National Center for Agricultural Utilization Research, 2Mycotoxin Prevention and Applied Microbiology Research Unit, National Center for Agricultural Utilization Research, 3Chemical Engineering and Material Science, Great Lakes Bioenergy Center, Michigan State University

Adaptive evolution and isolation techniques are described and demonstrated to yield derivatives of Scheffersomyces stipitis strain NRRL Y-7124 that are able to rapidly consume hexose and pentose mixed sugars in enzyme saccharified undetoxified hydrolyzates and to accumulate over 40 g/L ethanol.

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JoVE Journal

Gap Junctional Intercellular Communication: A Functional Biomarker to Assess Adverse Effects of Toxicants and Toxins, and Health Benefits of Natural Products
Brad L. Upham 1, Iva Sovadinová 2, Pavel Babica 2
1Department of Pediatrics & Human Development, Institute for Integrative Toxicology, Michigan State University, 2RECETOX — Research Centre for Toxic Compounds in the Environment, Faculty of Science, Masaryk University

This protocol describes a scalpel loading-fluorescent dye transfer technique that measures intercellular communication through gap junction channels. Gap junctional intercellular communication is a major cellular process by which tissue homeostasis is maintained and disruption of this cell signaling has adverse health effects.

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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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Environment

Building Double-decker Traps for Early Detection of Emerald Ash Borer
Deborah G. McCullough 1, Therese M. Poland 2
1Department of Entomology and Department of Forestry, Michigan State University, 2Northern Research Station, USDA Forest Service

Effective traps to attract and capture the emerald ash borer (EAB) are a key element of detecting and managing this invasive pest. Double-decker traps, placed in full sun near ash trees, incorporate visual and olfactory cues and were more likely to capture EAB than other trap designs in field trials.

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Biochemistry

Multimer-PAGE: A Method for Capturing and Resolving Protein Complexes in Biological Samples
Tyler Rhinesmith 1, Bryan A. Killinger 2, Akhil Sharma 3, Anna Moszczynska 3
1Physiology, Michigan State University, 2Center for Neurodegenerative Science Van Andel Institute, 3Pharmaceutical Sciences, Wayne State University

A method for stabilizing and separating native protein complexes from unmodified tissue lysate using an amine-reactive protein cross-linker coupled to a novel two-dimensional polyacrylamide gel electrophoresis (PAGE) system is presented.

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Genetics

Quantifying Abdominal Pigmentation in Drosophila melanogaster
Omid Saleh Ziabari 1, Alexander W. Shingleton 1
1Department of Biology, Lake Forest College

This work presents a method to quickly and precisely quantify the abdominal pigmentation of Drosophila melanogaster using digital image analysis. This method streamlines the procedures between phenotype acquisition and data analysis and includes specimen mounting, image acquisition, pixel value extraction, and trait measurement.

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Developmental Biology

Expansion and Adipogenesis Induction of Adipocyte Progenitors from Perivascular Adipose Tissue Isolated by Magnetic Activated Cell Sorting
Kyan Thelen 1, Nadia Ayala-Lopez 2, Stephanie W. Watts 2, G. Andres Contreras 1
1Department of Large Animal Clinical Sciences, Michigan State University, 2Department of Pharmacology and Toxicology, Michigan State University

Here we report a method for isolation of Adipocyte Progenitor Cell (APC) populations from Perivascular Adipose Tissue (PVAT) using Magnetic-activated Cell Sorting (MCS). This method allows for an increased isolation of APC per gram of adipose tissue when compared to Fluorescence-Activated Cell Sorting (FACS).

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Immunology and Infection

Come to the Light Side: In Vivo Monitoring of Pseudomonas aeruginosa Biofilm Infections in Chronic Wounds in a Diabetic Hairless Murine Model
Alessandra M. Agostinho Hunt 1, Jacob A. Gibson 1, Casandra L. Larrivee 1, Sandra O'Reilly 2, Svetlana Navitskaya 2, Julia V. Busik 2, Christopher M. Waters 1
1Department of Microbiology and Molecular Genetics, Michigan State University, 2Department of Physiology, Michigan State University

Here we describe a novel diabetic murine model utilizing hairless mice for real-time, non-invasive, monitoring of biofilm wound infections of bioluminescent Pseudomonas aeruginosa. This method can be adapted to evaluate infection of other bacterial species and genetically modified microorganisms, including multi-species biofilms, and test the efficacy of antibiofilm strategies.

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Behavior

How to Build a Dichoptic Presentation System That Includes an Eye Tracker
Cheng S. Qian 1, Jan W. Brascamp 1,2
1Department of Psychology, Michigan State University, 2Neuroscience Program, Michigan State University

We recently proposed a method that allows dichoptic visual stimulus presentation and binocular eye tracking simultaneously1. The key is the combination of an infrared eye tracker and the corresponding infrared-transparent mirrors. This manuscript provides an in depth protocol for initial setup and everyday operation.

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Bioengineering

Synthesis of Multi-walled Carbon Nanotubes Modified with Silver Nanoparticles and Evaluation of Their Antibacterial Activities and Cytotoxic Properties
Youngmin Seo *1, Chanhwi Park *2, Jaewoo Son 2, Kyungwoo Lee 2, Jangsun Hwang 2, Yeonho Jo 2, Dohyun Lee 2, Muhammad Saad Khan 2, Sachin Ganpat Chavan 2, Yonghyun Choi 2, Dasom Kim 2, Assaf A Gilad 3, Jonghoon Choi 2
1Center for Biomaterials, Biomedical Research Institute, Korea Institute of Science and Technology, 2School of Integrative Engineering, Chung-Ang University, 3Division of Synthetic Biology and Regenerative Medicine, Institute for Quantitative Health Science and Engineering, Michigan State University

In this study, antimicrobial nanomaterials were synthesized by acidic oxidation of multiwalled carbon nanotubes and subsequent reductive deposition of silver nanoparticles. Antimicrobial activity and cytotoxicity tests were performed with the as-prepared nanomaterials.

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Environment

Ammonia Fiber Expansion (AFEX) Pretreatment of Lignocellulosic Biomass
Shishir P. S. Chundawat 1, Ramendra K. Pal 1, Chao Zhao 1, Timothy Campbell 2, Farzaneh Teymouri 2, Josh Videto 2, Chandra Nielson 2, Bradley Wieferich 3, Leonardo Sousa 3, Bruce E. Dale 3, Venkatesh Balan 4, Sarvada Chipkar 5, Jacob Aguado 5, Emily Burke 5, Rebecca G. Ong 5
1Department of Chemical and Biochemical Engineering, Rutgers-State University of New Jersey, 2Michigan Biotechnology Institute (MBI), 3Department of Chemical Engineering and Materials Science, Michigan State University, 4Engineering Technology Department, Biotechnology Program, College of Technology, University of Houston, 5Department of Chemical Engineering, Michigan Technological University

Ammonia fiber expansion (AFEX) is a thermochemical pretreatment technology that can convert lignocellulosic biomass (e.g., corn stover, rice straw, and sugarcane bagasse) into a highly digestible feedstock for both biofuels and animal feed applications. Here, we describe a laboratory-scale method for conducting AFEX pretreatment on lignocellulosic biomass.

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Biology

Isolation of Enteric Glial Cells from the Submucosa and Lamina Propria of the Adult Mouse
Zhen Wang 1,4, Ramon Ocadiz-Ruiz 1, Sinju Sundaresan 1, Lin Ding 1, Michael Hayes 1, Nirakar Sahoo 3, Haoxing Xu 1,2, Juanita L. Merchant 1,2,5
1Department of Internal Medicine-Gastroenterology, University of Michigan, 2Department of Molecular and Integrative Physiology, University of Michigan, 3Department of Molecular, Cellular and Developmental Biology, University of Michigan, 4Department of Gastrointestinal Surgery, The First Affiliated Hospital of Guangxi Medical University, 5Division of Gastroenterology, University of Arizona College of Medicine

Here, we describe the isolation of enteric-glial cells from the intestinal-submucosa using sequential EDTA incubations to chelate divalent cations and then incubation in non-enzymatic cell recovery solution. Plating the resultant cell suspension on poly-D-lysine and laminin results in a highly enriched culture of submucosal glial cells for functional analysis.

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Biochemistry

The Identification of Sea Lamprey Pheromones Using Bioassay-Guided Fractionation
Anne M. Scott 1, Ke Li 1, Weiming Li 1
1Department of Fisheries and Wildlife, Michigan State University

Here, we present a protocol to isolate and characterize the structure, olfactory potency, and behavioral response of putative pheromone compounds of sea lampreys.

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Medicine

Fracture Apparatus Design and Protocol Optimization for Closed-stabilized Fractures in Rodents
Robert L Zondervan 1,2, Mitch Vorce 3, Nick Servadio 4, Kurt D. Hankenson 2
1College of Osteopathic Medicine, Michigan State University, 2Department of Orthopaedic Surgery, University of Michigan Medical School, 3Lymann Briggs College, Michigan State University, 4College of Engineering, Michigan State University

The goal of the protocol is to optimize the fracture generation parameters to yield consistent fractures. This protocol accounts for the variations in bone size and morphology that may exist between animals. Additionally, a cost-effective, adjustable fracture apparatus is described.

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Biology

Determination of Reproductive Competence by Confirming Pubertal Onset and Performing a Fertility Assay in Mice and Rats
Hanne M. Hoffmann 1
1Department of Animal Science, Michigan State University

Many treatments and genetic mutations impact the timing of sexual maturity and fertility. This protocol describes a non-invasive method to evaluate pubertal onset in mice and rats prior to setting up a fertility study in sexually mature animals.

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Environment

Separation of Spinach Thylakoid Protein Complexes by Native Green Gel Electrophoresis and Band Characterization using Time-Correlated Single Photon Counting
Eliezer Schwarz 1, Gary Blanchard 2
1Department of Plant Biology, Michigan State University, 2Department of Chemistry, Michigan State University

Here we present a protocol to separate solubilized thylakoid complexes by Native Green Gel electrophoresis. Green gel bands are subsequently characterized by Time Correlated Single Photon Counting (TCSPC) and basic steps for data analysis are provided.

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Chemistry

Large-scale Top-down Proteomics Using Capillary Zone Electrophoresis Tandem Mass Spectrometry
Elijah N. McCool *1, Rachele Lubeckyj *1, Xiaojing Shen 1, Qiang Kou 2, Xiaowen Liu 2,3, Liangliang Sun 1
1Department of Chemistry, Michigan State University, 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, 3Center for Computational Biology and Bioinformatics, Indiana University School of Medicine

A detailed protocol is described for the separation, identification, and characterization of proteoforms in protein samples using capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS). The protocol can be used for the high-resolution characterization of proteoforms in simple protein samples and the large-scale identification of proteoforms in complex proteome samples.

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Engineering

A Silicon-tipped Fiber-optic Sensing Platform with High Resolution and Fast Response
Guigen Liu *1, Qiwen Sheng *1, Weilin Hou 2, Matthew L. Reinke 3, Ming Han 1,4
1Department of Electrical and Computer Engineering, Michigan State University, 2Naval Research Laboratory, Stennis Space Center, 3Oak Ridge National Laboratory, 4Department of Electrical and Computer Engineering, University of Nebraska-Lincoln

This work reports an innovative silicon-tipped fiber-optic sensing platform (Si-FOSP) for high-resolution and fast-response measurement of a variety of physical parameters, such as temperature, flow, and radiation. Applications of this Si-FOSP span from oceanographic research, mechanical industry, to fusion energy research.

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Engineering

Evaluating Targeting Accuracy in the Focal Plane for an Ultrasound-guided High-intensity Focused Ultrasound Phased-array System
Ke Li 1,2, Jingfeng Bai 1,2, Yazhu Chen 1,2, Xiang Ji 1,2
1Biomedical Instrument Institute, School of Biomedical Engineering, Shanghai Jiao Tong University, 2Shanghai Med-X Engineering Center for Medical Equipment and Technology, Shanghai Jiao Tong University

This study describes a protocol to evaluate the targeting accuracy in the focal plane of an ultrasound-guided high-intensity focused ultrasound phased-array system.

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Environment

Measurement of Aerosols Optical Thickness of the Atmosphere using the GLOBE Handheld Sun Photometer
Maryssa Bradley 1, Morewell Gasseller 1
1Department of Physics, Xavier University of Louisiana

The goal of the methods presented here is to measure aerosol optical thickness of the atmosphere. The sun photometer is pointed at the sun and the largest voltage reading obtained on an in-built digital voltmeter is recorded. Atmospheric measurements such as barometric pressure and relative humidity are also performed.

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Biochemistry

Isolation of Lipoprotein Particles from Chicken Egg Yolk for the Study of Bacterial Pathogen Fatty Acid Incorporation into Membrane Phospholipids
Phillip C. Delekta 1, Todd A. Lydic 2, Neal D. Hammer 1
1Department of Microbiology and Molecular Genetics, Michigan State University, 2Department of Physiology, Michigan State University

This method provides a framework for studying incorporation of exogenous fatty acids from complex host sources into bacterial membranes, particularly Staphylococcus aureus. To achieve this, protocols for the enrichment of lipoprotein particles from chicken egg yolk and subsequent fatty acid profiling of bacterial phospholipids utilizing mass spectrometry are described.

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Neuroscience

Generation of Alpha-Synuclein Preformed Fibrils from Monomers and Use In Vivo
Joseph R. Patterson 1, Nicole K. Polinski 2, Megan F. Duffy 1,3, Christopher J. Kemp 1, Kelvin C. Luk 4, Laura A. Volpicelli-Daley 5, Nicholas M. Kanaan 1, Caryl E. Sortwell 1,3,6
1Department of Translational Science and Molecular Medicine, Michigan State University, 2The Michael J. Fox Foundation for Parkinson's Research, 3Neuroscience Program, Michigan State University, 4Center for Neurodegenerative Disease Research, Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine, 5Center for Neurodegeneration and Experimental Therapeutics, University of Alabama at Birmingham, 6Mercy Health Hauenstein Neuroscience Medical Center

The goal of this article is to outline the steps required for the generation of fibrils from monomeric alpha-synuclein, subsequent quality control, and use of the preformed fibrils in vivo.

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Chemistry

Enzymatic Synthesis of Epoxidized Metabolites of Docosahexaenoic, Eicosapentaenoic, and Arachidonic Acids
Joseph W. Woodman 1, Maris A. Cinelli 1, Amy Scharmen-Burgdolf 1, Kin Sing Stephen Lee 1
1Department of Pharmacology and Toxicology, Michigan State University

We present a method useful for large-scale enzymatic synthesis and purification of specific enantiomers and regioisomers of epoxides of arachidonic acid (AA), docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA) with the use of a bacterial cytochrome P450 enzyme (BM3).

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Biology

Silencing the Spark: CRISPR/Cas9 Genome Editing in Weakly Electric Fish
Savvas J. Constantinou 1, Linh Nguyen 2, Frank Kirschbaum 2, Vielka L. Salazar 3, Jason R. Gallant 1
1Department of Integrative Biology, Michigan State University, 2Faculty of Life Sciences, Unit of Biology and Ecology of Fishes, Humboldt University, 3Department of Biology, Cape Breton University

Here, a protocol is presented to produce and rear CRISPR/Cas9 genome knockout electric fish. Outlined in detail are the required molecular biology, breeding, and husbandry requirements for both a gymnotiform and a mormyrid, and injection techniques to produce Cas9-induced indel F0 larvae.

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Developmental Biology

Analysis of Congenital Heart Defects in Mouse Embryos Using Qualitative and Quantitative Histological Methods
Kristen Ball 1,2, Renee Kinne 2, Aitor Aguirre 1,2
1Department of Biomedical Engineering, Michigan State University, 2Institute for Quantitative Health Sciences and Engineering, Michigan State University

In this protocol, we describe procedures to qualitatively and quantitatively analyze developmental phenotypes in mice associated with congenital heart defects.

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JoVE Core

Flow Cytometry Analysis of Immune Cell Subsets within the Murine Spleen, Bone Marrow, Lymph Nodes and Synovial Tissue in an Osteoarthritis Model
Patrick Haubruck 1,2, Aimee C. Colbath 1,3, Yolanda Liu 1, Shihani Stoner 1, Cindy Shu 1, Christopher B. Little 1
1Raymond Purves Bone and Joint Research Laboratory, Institute of Bone and Joint Research, Kolling Institute, Royal North Shore Hospital, University of Sydney, 2HTRG - Heidelberg Trauma Research Group, Center for Orthopedics, Trauma Surgery and Spinal Cord Injury, Trauma and Reconstructive Surgery, Heidelberg University Hospital, 3Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University

Here, we describe a detailed and reproducible flow cytometry protocol to identify monocyte/macrophage and T-cell subsets using both extra- and intracellular staining assays within the murine spleen, bone marrow, lymph nodes and synovial tissue, utilizing an established surgical model of murine osteoarthritis.

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JoVE Core

Setup and Execution of the Rapid Cycle Deliberate Practice Death Notification Curriculum
Patrick G. Hughes 1, Kate E. Hughes 2, Mary J. Hughes 3,4, Lindsay Weaver 4, Lauren E. Falvo 4, Anna M. Bona 4, Dylan Cooper 4, Cherri Hobgood 4, Rami A. Ahmed 4
1Florida Atlantic University Schmidt College of Medicine, 2Department of Emergency Medicine, University of Arizona College of Medicine, 3Department of Osteopathic Medical Specialties, College of Osteopathic Medicine, Michigan State University, 4Indiana University School of Medicine

The goal is to demonstrate how to apply the rapid cycle deliberate practice debriefing technique to the GRIEV_ING death notification curriculum.

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Neuroscience

Subretinal Transplantation of Human Embryonic Stem Cell-Derived Retinal Tissue in a Feline Large Animal Model
Laurence M. Occelli *1, Felipe Marinho *1, Ratnesh K. Singh 2, Francois Binette 2, Igor O. Nasonkin 2, Simon M. Petersen-Jones 1
1College of Veterinary Medicine, Department of Small Animal Clinical Sciences, Michigan State University, 2Lineage Cell Therapeutics, Inc.

Presented here is a surgical technique for transplanting human pluripotent stem cell (hPSC)-derived retinal tissue into the subretinal space of a large animal model.

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Biochemistry

Complementation of Splicing Activity by a Galectin-3 - U1 snRNP Complex on Beads
Patricia G. Voss 1, Kevin C. Haudek 1, Ronald J. Patterson 2, John L. Wang 1
1Department of Biochemistry and Molecular Biology, Michigan State University, 2Department of Microbiology and Molecular Genetics, Michigan State University

This article describes the experimental procedures for (a) depletion of U1 snRNP from nuclear extracts, with concomitant loss of splicing activity; and (b) reconstitution of splicing activity in the U1-depleted extract by galectin-3 - U1 snRNP particles bound to beads covalently coupled with anti-galectin-3 antibodies.

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Neuroscience

Stereotaxic Intracranial Delivery of Chemicals, Proteins or Viral Vectors to Study Parkinson's Disease
Natalie Landeck 1, Melissa Conti Mazza 1, Megan Duffy 1, Christopher Bishop 2, Caryl E. Sortwell 3, Mark R. Cookson 1
1Cell Biology and Gene Expression Section, National Institute on Aging, National Institutes of Health, 2Behavioral Neuroscience Program, Department of Psychology, Binghamton University, 3Department of Translational Neuroscience, Mercy Health Hauenstein Neuroscience Medical Center, Michigan State University

We describe how to successfully inject solutions into specific brain areas of rodents using a stereotaxic frame. This survival surgery is a well-established method used to mimic various aspects of Parkinson's disease.

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Bioengineering

Establishing an Octopus Ecosystem for Biomedical and Bioengineering Research
Tyler VanBuren 1,2, Carolina Cywiak 1,2, Petra Telgkamp 1,2, Christiane L. Mallett 3,4, Galit Pelled 1,2,4
1Department of Biomedical Engineering, Michigan State University, 2Neuroengineering division, The Institute for Quantitative Health Science and Engineering, Michigan State University, 3Biomedical Imaging division, The Institute for Quantitative Health Science and Engineering, Michigan State University, 4Department of Radiology, Michigan State University

Understanding the unique physiological and anatomical structures of octopuses can greatly impact biomedical research. This guide demonstrates how to set-up and maintain a marine environment to accommodate this species and includes state-of-the-art imaging and analytical approaches to visualize octopus' nervous system anatomy and function.

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Neuroscience

Induction and Assessment of Levodopa-induced Dyskinesias in a Rat Model of Parkinson's Disease
Margaret E. Caulfield 1, Jennifer A. Stancati 1, Kathy Steece-Collier 1,2
1Department of Translational Neuroscience, College of Human Medicine, Michigan State University, 2Hauenstein Neuroscience Center, Mercy Health Saint Mary's

This article describes methods to induce and evaluate levodopa-induced dyskinesias in a rat model of Parkinson's disease. The protocol offers detailed information regarding the intensity and frequency of a range of dyskinetic behaviors, both dystonic and hyperkinetic, providing a reliable tool to test treatments targeting this unmet medical need.

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Biochemistry

Generating Self-Assembling Human Heart Organoids Derived from Pluripotent Stem Cells
Yonatan R. Lewis-Israeli 1,2, Brett D. Volmert 1,2, Mitchell A. Gabalski 1,2, Amanda R. Huang 1,2, Aitor Aguirre 1,2
1Institute for Quantitative Health Science and Engineering, Division of Developmental and Stem Cell Biology, Michigan State University, 2Department of Biomedical Engineering, College of Engineering, Michigan State University

Here, we describe a protocol to create developmentally relevant human heart organoids (hHOs) efficiently using human pluripotent stem cells by self-organization. The protocol relies on the sequential activation of developmental cues and produces highly complex, functionally relevant human heart tissues.

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Cancer Research

Intraductal Delivery and X-ray Visualization of Ethanol-Based Ablative Solution for Prevention and Local Treatment of Breast Cancer in Mouse Models
Elizabeth Kenyon 1,2, Erin K. Zaluzec 1,3, Katherine Powell 1,2, Maximilian Volk 1,4, Shatadru Chakravarty 2,5, Jeremy Hix 2,6, Ripla Arora 7,8, Jennifer J. Westerhuis 9,10, Matti Kiupel 11, Erik M. Shapiro 2,6, Lorenzo F. Sempere 1,2
1Precision Health Program, Michigan State University, 2Department of Radiology, College of Human Medicine, Michigan State University, 3Department of Pharmacology & Toxicology, College of Veterinary Medicine, Michigan State University, 4College of Osteopathic Medicine, Michigan State University, 5Advanced Materials Characterization Laboratory/Materials Research Center, Missouri University of Science and Technology, 6Institute for Quantitative (IQ) Health Science and Engineering Advanced Molecular Imaging Facility, Michigan State University, 7Department of Obstetrics Gynecology and Reproductive Biology, College of Human Medicine, Michigan State University, 8Institute for Quantitative (IQ) Health Science and Engineering, Michigan State University, 9Van Andel Research Institute, 10Miltenyi Biotec, 11Veterinary Diagnostic Laboratory, College of Veterinary Medicine, Michigan State University

A method of intraductal injection of reagents for an ethanol-based ablative solution to the mouse mammary ductal tree for in vivo imaging and breast cancer prevention is described. Injection directly into the nipple opening allows for targeting mammary epithelial cells with minimal collateral tissue damage.

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Genetics

Large-Scale SARS-CoV-2 Testing Utilizing Saliva and Transposition Sample Pooling
Joseph R. Patterson 1,2,3, Allyson Cole-Strauss 1,2,3, Nathan Kuhn 1,2, Carlene Mercier 3,4, Joseph Kochmanski 1,2, John A. Gerlach 5, Rhiannon M. LeVeque 3,4, Kerri A. Neugebauer 3,4,6, Kayla N. Conner 3,4, Jasper Gomez 3,4, Mark G. Hennes 3,4, Kaje’ne E. Thompson 3,4, Destinee L. Rytlewski 3,4, Chloe C. Bigwood 3,4, Amy Scharmen 3,4, Gabriel Simjanovski 3,4, Cassidy Riley 3,4, Jessica Donaldson 3,4, Dilann Yasin 3,4, Najwa Kouja 3,5, Zaria Contejean 3,4, Michaela Burnett 3,4, Shakhlo Aminova 3,4, Nat Ato Yawson 3,4,7, Simran B. Singh 3,4, Osama M. Alian 3,4, Carson D. Broeker 3,4,8, Erin K. Zaluzec 3,4,7, Morgan ONeill 3,4,9, Birgit Puschner 10, Aron Sousa 2, Laura Bix 11, Brian Jespersen 2, Claudia Holzman 12, Jade Mitchell 13, Ryan Julien 13, Yesim Askin 14, Danielle Barnes 14, Purna Durshanpalli 14, Doug Krum 14, Rett Weber 14, Morgan Patterson 14, Becky Anderson 14, Ryan Hunt 14, Benjamin O’Brien 14, Andrew Umstead 1,2, John S. Beck 1,2, Irving E. Vega 1,2, Caryl E. Sortwell 1,2, Jack W. Lipton 1,2
1Department of Translational Neuroscience, Michigan State University, 2College of Human Medicine, Michigan State University, 3COVID-19 Early Detection Program, Michigan State University, 4Department of Microbiology and Molecular Genetics, Michigan State University, 5Biomedical Laboratory Diagnostics Program, Michigan State University, 6Department of Plant, Soil, and Microbial Sciences, Michigan State University, 7Department of Pharmacology and Toxicology, Michigan State University, 8Department of Biochemistry and Molecular Biology, Michigan State University, 9College of Osteopathic Medicine, Michigan State University, 10College of Veterinary Medicine, Michigan State University, 11School of Packaging, Michigan State University, 12Department of Epidemiology and Biostatistics, Michigan State University, 13Department of Biosystems Engineering, Michigan State University, 14Enterprise Services, Michigan State University

The protocol is intended to serve as a blueprint for universities and other organizations considering large-scale testing for SARS-CoV-2 or developing preparedness plans for future viral outbreaks.

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Cancer Research

X-Ray Visualization of Intraductal Ethanol-Based Ablative Treatment for Prevention of Breast Cancer in Rat Models
Elizabeth Kenyon 1,2, Erin Zaluzec 1,3, Katherine Powell 1,2, Maximilian Volk 1,4, Shatadru Chakravarty 2,5, Jeremy Hix 2,6, Matti Kiupel 7, Erik M. Shapiro 2, Lorenzo F. Sempere 1,2
1Precision Health Program, Michigan State University, 2Department of Radiology, Michigan State University, 3Department of Pharmacology & Toxicology, Michigan State University, 4College of Osteopathic Medicine, Michigan State University, 5TechInsights Inc., 6IQ Advanced Molecular Imaging Facility, Michigan State University, 7Veterinary Diagnostic Laboratory, College of Veterinary Medicine, Michigan State University

A procedure for the delivery of a chemical ablative solution to the rat mammary ductal tree for image-guided preventive treatment of breast cancer is described. Mammary epithelial cells can be targeted with minimal collateral tissue damage through cannulation directly into the nipple opening and intraductal infusion of a 70% ethanol-based ablative solution.

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Medicine

Establishment of a Simple and Effective Rat Model for Intraoperative Parathyroid Gland Imaging
Fan Chen *1,2, Chun Liu *3, Peng Guo 3, Weihui Zheng 1,2
1Key Laboratory of Head & Neck Cancer Translational Research of Zhejiang Province, Zhejiang Cancer Hospital, 2The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), 3Institute of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences

To date, the development of parathyroid gland (PG) identification methods is limited by the lack of animal models in preclinical research. Here, we establish a simple and effective rat model for intraoperative PG imaging and evaluate its effectiveness by using iron oxide nanoparticles as a novel PG contrast agent.

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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.

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Biology

Plastoglobule Lipid Droplet Isolation from Plant Leaf Tissue and Cyanobacteria
Kiran-Kumar Shivaiah 1,2, Febri A. Susanto 1,2, Elsinraju Devadasu 1,2, Peter K. Lundquist 1,2
1Department of Biochemistry and Molecular Biology, Michigan State University, 2Plant Resilience Institute, Michigan State University

A fast and efficient protocol is presented for the isolation of plastoglobule lipid droplets associated with various photosynthetic organisms. The successful preparation of isolated plastoglobules is a crucial first step that precedes detailed molecular investigations such as proteomic and lipidomic analyses.

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Genetics

Daily Transfers, Archiving Populations, and Measuring Fitness in the Long-Term Evolution Experiment with Escherichia coli
Jeffrey E. Barrick 1,2, Zachary D. Blount 2,3, Devin M. Lake 2,4, Jack H. Dwenger 1, Jesus E. Chavarria-Palma 1, Minako Izutsu 2,3, Michael J. Wiser 2,5
1Department of Molecular Biosciences, The University of Texas at Austin, 2BEACON Center for the Study of Evolution in Action, Michigan State University, 3Department of Microbiology and Molecular Genetics, Michigan State University, 4Department of Integrative Biology, Michigan State University, 5Biological Sciences Program, Michigan State University

This protocol describes how to maintain the Escherichia coli Long-Term Evolution Experiment (LTEE) by performing its daily transfers and periodic freeze-downs and how to conduct competition assays to measure fitness improvements in evolved bacteria. These procedures can serve as a template for researchers starting their own microbial evolution experiments.

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Bioengineering

Multimodal Cross-Device and Marker-Free Co-Registration of Preclinical Imaging Modalities
Mirko Thamm 1,2, Justin J. Jeffery 3, Yapei Zhang 4, Bryan R. Smith 4, Stephen Marchant 5, Fabian Kiessling 1,6, Felix Gremse 1,2
1Institute for Experimental Molecular Imaging, RWTH Aachen University, 2Gremse-IT GmbH, 3Carbone Cancer Center, University of Wisconsin, 4Department of Biomedical Engineering and Institute for Quantitative Health Science and Engineering, Michigan State University, 5MediLumine Inc., 6Fraunhofer Institute for Digital Medicine MEVIS

The combination of multiple imaging modalities is often necessary to gain a comprehensive understanding of pathophysiology. This approach utilizes phantoms to generate a differential transformation between the coordinate systems of two modalities, which is then applied for co-registration. This method eliminates the need for fiducials in production scans.

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Neuroscience

Using Live-Cell Imaging to Measure the Effects of Pathological Proteins on Axonal Transport in Primary Hippocampal Neurons
Rebecca L. Mueller 1,2, Nicholas M. Kanaan 1,2,3, Benjamin Combs 1
1Department of Translational Neuroscience, College of Human Medicine, Michigan State University, 2Neuroscience Program, Michigan State University, 3Hauenstein Neuroscience Center, Mercy Health Saint Mary’s

Here, we demonstrate how to combine transfection of primary hippocampal rodent neurons with live-cell confocal imaging to analyze pathological protein-induced effects on axonal transport and identify mechanistic pathways mediating these effects.

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Cancer Research

Monitoring of Nanodrug Accumulation in Murine Breast Cancer Metastases
Alan Halim 1, Sujan Kumar Mondal 1,2, Nasreen Al-Qadi 1, Elizabeth Kenyon 1,2, Keith MacRenaris 3,5, Thomas V. O’Halloran 4,5, Zdravka Medarova 6, Anna Moore 1,2
1Precision Health Program, Michigan State University, 2Department of Radiology, College of Human Medicine, Michigan State University, 3Department of Microbiology, Genetics, & Immunology, College of Natural Science, Michigan State University, 4Department of Chemistry, College of Natural Science, Michigan State University, 5Elemental Health Institute, Michigan State University, 6Transcode Therapeutics Inc.

Here, we describe the protocol for in vivo delivery of magnetic iron oxide nanoparticles carrying RNA oligomers to metastatic breast cancer in animal models, providing a clinically viable approach for the therapeutic silencing of oncogenic nucleic acids.

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Bioengineering

Porous Substrate-Based Electroporation with Transepithelial Electrical Impedance Monitoring
Sawyer R. Lorenzen 1,2, Justin R. Brooks 1, Tyler C. Heiman 1, Ruiguo Yang 1,2,3
1Department of Mechanical and Materials Engineering, University of Nebraska-Lincoln, 2Department of Biomedical Engineering, Michigan State University, 3Institute for Quantitative Health Science and Engineering (IQ), Michigan State University

Porous substrate electroporation (PSEP) pairs consistent, high throughput delivery with high cell viability. Introduction of transepithelial electrical impedance (TEEI) measurements provides insight into the intermediate processes of PSEP and allows for label-free delivery. This article discusses a method for performing PSEP delivery experiments and TEEI measurement analysis simultaneously.

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Nanoparticle Delivery of an Oligonucleotide Payload in a Glioblastoma Multiforme Animal Model

Nanoparticle Delivery of an Oligonucleotide Payload in a Glioblastoma Multiforme Animal Model
Bryan D. Kim 1,2, Sujan K. Mondal 1,3, Elizabeth Kenyon 1,3, Ming Chen 1,3, Christiane L. Mallett 3,4, Ana C. deCarvalho 5, Zdravka Medarova 6, Anna Moore 1,3
1Precision Health Program, Michigan State University, 2Department of Biomedical Engineering, College of Engineering, Michigan State University, 3Department of Radiology, College of Human Medicine, Michigan State University, 4Institute for Quantitative Health Science and Engineering, Michigan State University, 5Henry Ford Health, 6Transcode Therapeutics Inc.

The blood-brain barrier is a significant hurdle in the delivery of therapies for glioblastoma, a disease for which there is no cure. Here, we report an in vivo image-guided iron oxide therapeutic nano platform that can bypass this physiological barrier by virtue of size and accumulate in the tumor.

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Biochemistry

Robust Mitochondrial Isolation from Rodent Cardiac Tissue
Alyssa C. Vadovsky 1, Melissa Quinn 2, Tianyi Xia 1, Yan Levitsky 1, Jason N. Bazil 1
1Department of Physiology, Michigan State University, 2Department of Kinesiology, Michigan State University

Bioenergetic and metabolomic studies on mitochondria have revealed their multifaceted role in many diseases, but the isolation methods for these organelles vary. The method detailed here is capable of purifying high-quality mitochondria from multiple tissue sources. Quality is determined by respiratory control ratios and other metrics assessed with high-resolution respirometry.

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