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30 ARTICLES PUBLISHED IN JoVE

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Immunology and Infection

Finger-stick Blood Sampling Methodology for the Determination of Exercise-induced Lymphocyte Apoptosis
James Navalta 1, Brian McFarlin *2, Richard Simpson 2, Elizabeth Fedor 1, Holly Kell 1, Scott Lyons 1, Scott Arnett 1, Mark Schafer 1
1Department of Kinesiology, Recreation, and Sport, Western Kentucky University, 2Department of Health and Human Performance, University of Houston

Exercise is capable of inducing apoptosis in immune cells. There are various measurement limitations, particularly relating to the amount of time required to isolate and treat a blood sample prior to the assessment. Demonstrated is a rapid and minimally invasive procedure for the analysis of exercise-induced lymphocyte apoptosis.

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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

Specimen Preparation, Imaging, and Analysis Protocols for Knife-edge Scanning Microscopy
Yoonsuck Choe 1, David Mayerich 2, Jaerock Kwon 3, Daniel E. Miller 1, Chul Sung 1, Ji Ryang Chung 1, Todd Huffman 4, John Keyser 1, Louise C. Abbott 5
1Department of Computer Science and Engineering, Texas A&M University, 2Beckman Institute for Advanced Science and Technology, University of Illinois, 3Department of Electrical and Computer Engineering, Kettering University, 43Scan, 5Department of Veterinary Integrative Biosciences, Texas A&M University

The full process from brain specimen preparation to serial sectioning imaging using the Knife-Edge Scanning Microscope, to data visualization and analysis is described. This technique is currently used to acquire mouse brain data, but it is applicable to other organs, other species.

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Neuroscience

The FlyBar: Administering Alcohol to Flies
Kim van der Linde 1, Emiliano Fumagalli 2, Gregg Roman 2, Lisa C. Lyons 1
1Department of Biological Science, Florida State University, 2Department of Biology and Biochemistry, Biology of Behavior Institute, University of Houston

Drosophila has emerged as a significant model system for dissecting the cellular and molecular underpinnings of behavioral responses to alcohol. Here we present a protocol for the collection of alcohol sensitivity data in a circadian context that can be easily applied to other experiments and is well-suited for undergraduate research.

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Behavior

Simultaneous Scalp Electroencephalography (EEG), Electromyography (EMG), and Whole-body Segmental Inertial Recording for Multi-modal Neural Decoding
Thomas C. Bulea 1,2, Atilla Kilicarslan 2, Recep Ozdemir 2,3,4, William H. Paloski 3,4, Jose L. Contreras-Vidal 2,4,5
1Functional and Applied Biomechanics Group, National Institutes of Health, 2Laboratory for Non-invasive Brain-Machine Interface Systems, Department of Electrical and Computer Engineering, University of Houston, 3Department of Health and Human Performance, University of Houston, 4Center for Neuromotor & Biomechanics Research, University of Houston, 5Department of Biomedical Engineering, University of Houston

Development of an effective brain-machine-interface (BMI) system for restoration and rehabilitation of bipedal locomotion requires accurate decoding of user's intent. Here we present a novel experimental protocol and data collection technique for simultaneous non-invasive acquisition of neural activity, muscle activity, and whole-body kinematics during various locomotion tasks and conditions.

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JoVE Core

Qualitative Identification of Carboxylic Acids, Boronic Acids, and Amines Using Cruciform Fluorophores
Thimon Schwaebel 1, Rio Carlo Lirag 2, Evan A. Davey 1, Jaebum Lim 2, Uwe H. F. Bunz 1, Ognjen Š. Miljanić 2
1Organisch-Chemisches Institut, Ruprecht-Karls-Universität Heidelberg, 2Department of Chemistry, University of Houston

Cross-conjugated cruciform fluorophores based on 1,4-distyryl-2,5-bis(arylethynyl)benzene and benzobisoxazole nuclei can be used to qualitatively identify diverse Lewis acidic and Lewis basic analytes. This method relies on the differences in emission colors of the cruciforms that are observed upon analyte addition. Structurally closely related species can be distinguished from each other.

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Medicine

Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells
Anne Katchy 1, Cecilia Williams 1
1Center for Nuclear Receptors and Cell Signaling, Department of Biology and Biochemistry, University of Houston

Molecular signaling through both estrogen and microRNAs are critical in breast cancer development and growth. Estrogen activates the estrogen receptors, which are transcription factors. Many transcription factors can regulate the expression of microRNAs, and estrogen-regulated microRNAs can be profiled using different large-scale techniques.

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Environment

Tracking Microbial Contamination in Retail Environments Using Fluorescent Powder - A Retail Delicatessen Environment Example
Sujata A. Sirsat 1, Kawon Kim 1, Kristen E. Gibson 2, Phillip G. Crandall 2, Steven C. Ricke 2, Jack A. Neal 1
1Conrad N. Hilton College of Hotel and Restaurant Management, University of Houston, 2Center for Food Safety, Department of Food Science, University of Arkansas

The overall goal of this study was to demonstrate the potential cross contamination mechanism of foodborne pathogen Listeria monocytogenes in a retail deli setting. This methodology may be applied to a variety of different environments to track pathogen contamination.  

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Chemistry

Confocal Imaging of Confined Quiescent and Flowing Colloid-polymer Mixtures
Rahul Pandey *1, Melissa Spannuth *1, Jacinta C. Conrad 1
1Chemical and Biomolecular Engineering Department, University of Houston

Confocal microscopy is used to image quiescent and flowing colloid-polymer mixtures, which are studied as model systems for attractive suspensions. Image analysis algorithms are used to calculate structural and dynamic metrics for the colloidal particles that measure changes due to geometric confinement.

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Behavior

A Novel Experimental and Analytical Approach to the Multimodal Neural Decoding of Intent During Social Interaction in Freely-behaving Human Infants
Jesus G. Cruz-Garza 1, Zachery R. Hernandez 1, Teresa Tse 1,2,3, Eunice Caducoy 1,3, Berdakh Abibullaev 1, Jose L. Contreras-Vidal 1,2
1Laboratory for Noninvasive Brain-Machine Interface Systems, Department of Electrical and Computer Engineering, University of Houston, 2Department of Biomedical Engineering, University of Houston, 3Department of Biology and Biochemistry, University of Houston

This protocol presents a novel methodology for the neural decoding of intent from freely-behaving infants during unscripted social interaction with an actor. Neural activity is acquired using non-invasive high-density active scalp electroencephalography (EEG). Kinematic data is collected with inertial measurement units and supplemented with synchronized video recording.

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Medicine

Modeling Myotonic Dystrophy 1 in C2C12 Myoblast Cells
Rui Liang *1, Wei Dong *1,3, Xiaopeng Shen 1, Xiaoping Peng 1,3, Angie G. Aceves 1,2, Yu Liu 1
1Department of Biology and Biochemistry, University of Houston, 2Department of Economics, University of Houston, 3Department of Cardiology, The First Affiliated Hospital of Nanchang University

In this protocol, we present the procedures in establishing myotonic dystrophy 1 myoblast models, including optimized C2C12 cell maintenance, gene transfection/transduction, and myocyte differentiation.

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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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Bioengineering

Image-guided, Laser-based Fabrication of Vascular-derived Microfluidic Networks
Keely A. Heintz 1, David Mayerich 2, John H. Slater 1,3
1Department of Biomedical Engineering, University of Delaware, 2Department of Electrical and Computer Engineering, University of Houston, 3Delaware Biotechnology Institute

This protocol outlines the implementation of image-guided, laser-based hydrogel degradation to fabricate vascular-derived, biomimetic microfluidic networks embedded in poly(ethylene glycol) diacrylate (PEGDA) hydrogels. These biomimetic microfluidic systems may be useful for tissue engineering applications, generation of in vitro disease models, and fabrication of advanced "on-a-chip" devices.

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Immunology and Infection

A Protocol to Characterize the Morphological Changes of Clostridium difficile in Response to Antibiotic Treatment
Bradley Endres 1, Eugénie Bassères 1, Tasnuva Rashid 1, Long Chang 2, M. Jahangir Alam 1, Kevin W Garey 1
1Department of Pharmacy Practice and Translational Research, University of Houston College of Pharmacy, 2Department of Electrical and Computer Engineering, University of Houston

Antibiotic efficacy is most commonly determined by conducting killing kinetic studies and measuring colony forming units (CFUs). By integrating scanning electron microscopy (SEM) with these standard methods, we can distinguish the pharmacological effects of treatment between different antibiotics.

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Biology

Concurrent EEG and Functional MRI Recording and Integration Analysis for Dynamic Cortical Activity Imaging
Thinh Nguyen *1, Thomas Potter *1, Christof Karmonik 2, Robert Grossman 2, Yingchun Zhang 1,3
1Department of Biomedical Engineering, Cullen College of Engineering, University of Houston, 2Department of Neurosurgery, Houston Methodist Hospital and Research Institute, 3Guangdong Provincial Work Injury Rehabilitation Center

An EEG-fMRI multimodal imaging method, known as the spatiotemporal fMRI-constrained EEG source imaging method, is described here. The presented method employs conditionally-active fMRI sub-maps, or priors, to guide EEG source localization in a manner that improves spatial specificity and limits erroneous results.

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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

Pancreatic Islet Embedding for Paraffin Sections
Yahui Kong 1, Pantea Ebrahimpour 1,2, Yu Liu 3, Chaoxing Yang 1, Laura C. Alonso 1
1Diabetes Center of Excellence, UMass Medical School, 2Department of Medicine, Saint Vincent Hospital, 3Department of Pathology, Morphology Core, UMass Medical School

Ex vivo pancreatic islet studies are important for diabetes research. Existing techniques to study cultured islets in their native 3-dimensional architecture are time consuming, inefficient, and infrequently used. This work describes a new, simple, and efficient method for generating high-quality paraffin sections of whole cultured islets.

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Biochemistry

Dual DNA Rulers to Study the Mechanism of Ribosome Translocation with Single-Nucleotide Resolution
Heng Yin 1, Shoujun Xu 1, Yuhong Wang 2
1Department of Chemistry, University of Houston, 2Department of Biology and Biochemistry, University of Houston

Dual DNA ruler assay is developed to determine the mRNA position during ribosome translocation, which relies on the dissociation forces of the formed DNA-mRNA duplexes. With single-nucleotide resolution and capability of reaching both ends of mRNA, it can provide mechanistic insights for ribosome translocation and probe other nucleic acid displacements.

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JoVE Core

Quantification of Protein Interaction Network Dynamics using Multiplexed Co-Immunoprecipitation
Emily A. Brown 1,2, Steven C. Neier 3,4, Claudia Neuhauser 5, Adam G. Schrum 6,7,8, Stephen E.P. Smith 1,2,9
1Center for Integrative Brain Research, Seattle Children's Research Institute, 2Graduate Program in Neuroscience, University of Washington, 3Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Department of Medicine, Harvard Medical School, 4Broad Institute of Harvard and MIT, 5Department of Mathematics, University of Houston, 6Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, 7Department of Surgery, School of Medicine, University of Missouri, 8Department Bioengineering, College of Engineering, University of Missouri, 9Department of Pediatrics, University of Washington

Quantitative Multiplex Immunoprecipitation (QMI) uses flow cytometry for sensitive detection of differences in the abundance of targeted protein-protein interactions between two samples. QMI can be performed using a small amount of biomaterial, does not require genetically engineered tags, and can be adapted for any previously defined protein interaction network.

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Developmental Biology

A Layered Mounting Method for Extended Time-Lapse Confocal Microscopy of Whole Zebrafish Embryos
Sanat Upadhyay 1, Leoncio Vergara 2, Pranjali Shah 2, Jan-Åke Gustafsson 3,4, Ioannis Kakadiaris 1,4, Maria Bondesson 5
1Computational Biomedicine Lab, Texas Institute of Measurement Evaluation and Statistics, University of Houston, 2Institute of Biosciences and Technology, Texas A&M Health Science Center, 3Department of Biology and Biochemistry, Center for Nuclear Receptors and Cell Signaling, University of Houston, 4Department of Biosciences and Nutrition, Novum, Karolinska Institutet, 5Department of Intelligent Systems Engineering, Indiana University

This article describes a method to mount fragile zebrafish embryos for extended time-lapse confocal microscopy. This low-cost method is easy to perform using regular glass-bottom microscopy dishes for imaging on any inverted microscope. The mounting is performed in layers of agarose at different concentrations.

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Medicine

Traditional Trail Making Test Modified into Brand-new Assessment Tools: Digital and Walking Trail Making Test
Wei Wei *1, Hóngyi Zhào *1,2, Yu Liu 2, Yonghua Huang 1
1Department of Neurology, The Seventh Medical Center of PLA Army General Hospital, 2Department of Neurology, NO 984 Hospital of PLA

Here, we present a protocol to show how to perform two types of cognitive assessment tools derived from the paper-pencil version of the Trail Making Test.

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Engineering

Evaluation of an Exclusive Spur Dike U-Turn Design with Radar-Collected Data and Simulation
Yang Shao 1, Hongtao Yu 1, Huan Wu 2, Xueyan Han 1, Xizhen Zhou 1, Christian G. Claudel 3, Hualing Zhang 4, Chen Yang 5
1Highway Academy, Chang'an University, 2Human Resource Department, Xi'an Shiyou University, 3Cockrell School of Engineering, University of Texas at Austin, 4Department of Earth and Atmospheric Science, University of Houston, 5School of Foreign Languages, Xi'an Shiyou University

This protocol describes the process of solving a microscopic traffic problem with simulation. The whole process contains a detailed description of data collection, data analysis, simulation model build, simulation calibration, and sensitive analysis. Modifications and troubleshooting of the method are also discussed.

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Immunology and Infection

Visualization of Macrophage Lytic Cell Death During Mycobacterial Infection in Zebrafish Embryos via Intravital Microscopy
Liangfei Niu 1, Cong Wang 1, Kaile Zhang 1,2, Miaomiao Kang 1,2, Rui Liang 1, Xiaonan Zhang 1, Bo Yan 1
1Shanghai Public Health Clinical Center, Fudan University, 2School of Life Sciences, Bengbu Medical College

This protocol describes a technique for visualizing macrophage behavior and death in embryonic zebrafish during Mycobacterium marinum infection. Steps for the preparation of bacteria, infection of the embryos, and intravital microscopy are included. This technique may be applied to the observation of cellular behavior and death in similar scenarios involving infection or sterile inflammation.

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Immunology and Infection

High-throughput Screening of Chemical Compounds to Elucidate Their Effects on Bacterial Persistence
Prashant Karki 1, Mehmet A. Orman 1
1Department of Chemical and Biomolecular Engineering, University of Houston

In this method paper, we present a high-throughput screening strategy to identify chemical compounds, such as osmolytes, that have a significant impact on bacterial persistence.

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Medicine

Digital Handwriting Analysis of Characters in Chinese Patients with Mild Cognitive Impairment
Hóngyi Zhào *1,2, Yanhai Zhang *3, Cuiqiao Xia 1, Yu Liu 2, Zhiyi Li 2, Yonghua Huang 1
1Department of Neurology, Chinese PLA General Hospital, 2Department of Neurology, NO 984 Hospital of PLA, 3Department of Cadre Ward, Xijing 986 Hospital, Fourth Military Medical University

This study proposed a digital handwriting analysis of characters in individuals with mild cognitive impairment to find more information than is revealed by traditional pencil–paper handwriting analysis.

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Cancer Research

Characterization and Functional Prediction of Bacteria in Ovarian Tissues
Lanbo Zhao *1, Weichu Zhao *2, Qi Wang *1, Dongxin Liang 1, Yu Liu 3, Guoxing Fu 4, Lu Han 1, Yiran Wang 1, Chao Sun 1, Qing Wang 1, Qing Song 1, Qiling Li 1, Qinrui Lu 1
1Department of Obstetrics and Gynecology, First Affiliated Hospital of Xi’an Jiaotong University, 2School of Social Sciences, University of Manchester, 3Department of Pathology, First Affiliated Hospital of Xi’an Jiaotong University, 4Omega Bioservices Inc

Immunohistochemistry staining and 16S ribosomal RNA gene (16S rRNA gene) sequencing were performed in order to discover and distinguish bacteria in cancerous and noncancerous ovarian tissues in situ. The compositional and functional differences of the bacteria were predicted by using BugBase and Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt).

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Biology

Serial Block-Face Scanning Electron Microscopy (SBF-SEM) of Biological Tissue Samples
Justin A. Courson 1, Paul T. Landry 1, Thao Do 1, Eric Spehlmann 2, Pascal J. Lafontant 2, Nimesh Patel 1, Rolando E. Rumbaut 3,4, Alan R. Burns 1,4
1College of Optometry, University of Houston, 2Department of Biology, DePauw University, 3Center for Translational Research on Inflammatory Diseases (CTRID), Michael E. DeBakey Veterans Affairs Medical Center, 4Children’s Nutrition Center, Baylor College of Medicine

This protocol outlines a routine method for using serial block-face scanning electron microscopy (SBF-SEM), a powerful 3D imaging technique. Successful application of SBF-SEM hinges on proper fixation and tissue staining techniques, as well as careful consideration of imaging settings. This protocol contains practical considerations for the entirety of this process.

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JoVE Journal

Single Molecule Fluorescence Energy Transfer Study of Ribosome Protein Synthesis
Yuhong Wang 1
1Department of Biology and Biochemistry, University of Houston

Single molecule fluorescence energy transfer is a method that tracks the tRNA dynamics during ribosomal protein synthesis. By tracking individual ribosomes, inhomogeneous populations are identified, which shed light on mechanisms. This method can be used to track biological conformational changes in general to reveal dynamic-function relationships in many other complexed biosystems. Single molecule methods can observe non-rate limiting steps and low-populated key intermediates, which are not accessible by conventional ensemble methods due to the average effect.

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Medicine

An Epithelial Abrasion Model for Studying Corneal Wound Healing
Prince K. Akowuah 1, Angie De La Cruz 1, C. Wayne Smith 2, Rolando E. Rumbaut 2,3, Alan R. Burns 1,2
1College of Optometry, University of Houston, 2Children’s Nutrition Research Center, Baylor College of Medicine, 3Center for Translational Research on Inflammatory Diseases (CTRID), Michael E. DeBakey Veterans Affairs Medical Center

Here, a protocol for creating a central corneal epithelial abrasion wound in the mouse using a trephine and a blunt golf club spud is described. This corneal wound healing model is highly reproducible and is now being used to evaluate compromised corneal wound healing in the context of diseases.

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Behavior

Physical Activity Measurement in Children Accepting Table Tennis Training
Xin Zhang *1, Cuiqiao Xia *2, Xiuli Zhao 3, Yu Liu 3, Hóngyi Zhào 2,3, Yonghua Huang 2
1Department of Sport, Inner Mongolia Medical University, 2Department of Neurology, The Seventh Medical Center of PLA General Hospital, 3Department of Neurology, NO 984 Hospital of PLA

This study proposes an accelerometer-based method to objectively measure physical activity (PA) and leisure time physical activity (LTPA) in Chinese children accepting table tennis training in clubs.

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