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34 ARTICLES PUBLISHED IN JoVE

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Biology

Obtaining Hemocytes from the Hawaiian Bobtail Squid Euprymna scolopes and Observing their Adherence to Symbiotic and Non-Symbiotic Bacteria
Andrew J. Collins 1, Spencer V. Nyholm 1
1Department of Molecular and Cell Biology, University of Connecticut

This video will demonstrate how to obtain hemocytes (blood cells) from the Hawaiian bobtail squid, Euprymna scolopes for use in cell biological and bacterial adhesion assays. Hemocytes will be stained with a fluorescent dye and exposed to GFP-labeled bacteria.

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Biology

Window on a Microworld: Simple Microfluidic Systems for Studying Microbial Transport in Porous Media
Dmitry A. Markov 1,2, Philip C. Samson 1, David K. Schaffer 1, Adit Dhummakupt 1, John P. Wikswo 1,2,3,4, Leslie M. Shor 5,6
1Vanderbilt Institute for Integrative Biosystems Research and Education, Vanderbilt University, 2Department of Biomedical Engineering, Vanderbilt University, 3Department of Molecular Physiology and Biophysics, Vanderbilt University, 4Department of Physics and Astronomy, Vanderbilt University, 5Department of Chemical, Materials and Biomolecular Engineering, University of Connecticut, 6Center for Environmental Sciences and Engineering, University of Connecticut

Microfluidic devices can be used to visualize complex natural processes in real time and at the appropriate physical scales. We have developed a simple microfluidic device that mimics key features of natural porous media for studying growth and transport of bacteria in the subsurface.

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Neuroscience

In utero Electroporation followed by Primary Neuronal Culture for Studying Gene Function in Subset of Cortical Neurons
Heather Rice 1, Seiyam Suth 1, William Cavanaugh 1, Jilin Bai 2, Tracy L. Young-Pearse 1
1Center for Neurologic Diseases, Brigham and Woman's Hospital and Harvard Medical School, 2Department of Physiology and Neurobiology, University of Connecticut

In utero electroporation is a valuable method for transfecting neuronal progenitor cells in vivo. Depending upon the placement of the electrodes and the developmental timepoint of electroporation, certain subsets of cortical cells can be targeted. Targeted cells can then be analyzed in vivo or in vitro for effects of genetic alteration.

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Biology

Detection of Post-translational Modifications on Native Intact Nucleosomes by ELISA
Bo Dai 1, Farida Dahmani 2, Joseph A. Cichocki 3, Lindsey C. Swanson 2, Theodore P. Rasmussen 3
1Institute for Stem Cell Biology and Regenerative Medicine, Stanford University , 2Department of Molecular and Cell Biology, University of Connecticut, 3Department of Pharmaceutical Sciences, University of Connecticut

Nucleosome ELISA (NU-ELISA) is a sensitive and quantitative method to detect global patterns of post-translational modifications in preparations of native, intact nucleosomes. These modifications include methylations, acetylations, and phosphorylations at specific histone amino acid residues, and hence NU-ELISA provides a global proteomic assay of the overall chromatin modification states of specific cell types.

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Biology

Insulin Injection and Hemolymph Extraction to Measure Insulin Sensitivity in Adult Drosophila melanogaster
Aaron T. Haselton 1, Yih-Woei C. Fridell 2
1Department of Biology, State University of New York, 2Allied Health Sciences, University of Connecticut

Conserved insulin signaling pathways found in the fruit fly Drosophila melanogaster make this organism a potential tool for modeling metabolic disorders including type II diabetes. To this end, it is critical to establish physiological assays to effectively measure systemic insulin action in peripheral glucose disposal in the adult fly.

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Biology

Measurement of Cellular Chemotaxis with ECIS/Taxis
Kathryn M. Pietrosimone 1, Xiuyin Yin 2, David A. Knecht 1, Michael A. Lynes 1
1Molecular and Cell Biology, University of Connecticut, 2University of Connecticut

The ECIS/Taxis system is an automated, real-time assay that measures cellular chemotaxis. In this assay, cells move beneath a layer of agarose to arrive at a target electrode. Cellular movement is measured by the onset of resistance to AC current 0.

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Medicine

Portable Intermodal Preferential Looking (IPL): Investigating Language Comprehension in Typically Developing Toddlers and Young Children with Autism
Letitia R. Naigles 1, Andrea T. Tovar 1
1Department of Psychology, University of Connecticut

A reliable home-based way to assess the language comprehension of very young typically developing children, as well as those with autism, is described. The method analyzes children's eye gaze while viewing side-by-side images but hearing an audio that matches only one image. Stimuli are designed with young participants in mind.

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Engineering

Tensile Strength of Resorbable Biomaterials
Peiman Shahbeigi-Roodposhti 1, Sina Shahbazmohamadi 1
1University of Connecticut

Tensile Strength of Resorbable Biomaterials

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Engineering

Biodistribution of Nano-drug Carriers: Applications of SEM
Peiman Shahbeigi-Roodposhti 1, Sina Shahbazmohamadi 1
1University of Connecticut

Biodistribution of Nano-drug Carriers: Applications of SEM

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Engineering

Acquisition and Analysis of an ECG (electrocardiography) Signal
Peiman Shahbeigi-Roodposhti 1, Sina Shahbazmohamadi 1
1University of Connecticut

Acquisition and Analysis of an ECG (electrocardiography) Signal

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Engineering

Micro-CT Imaging of a Mouse Spinal Cord
Peiman Shahbeigi-Roodposhti 1, Sina Shahbazmohamadi 1
1University of Connecticut

Micro-CT Imaging of a Mouse Spinal Cord

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Engineering

Imaging Biological Samples with Optical and Confocal Microscopy
Peiman Shahbeigi-Roodposhti 1, Sina Shahbazmohamadi 1
1University of Connecticut

Imaging Biological Samples with Optical and Confocal Microscopy

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Engineering

Ceramic-matrix Composite Materials and Their Bending Properties
Sina Shahbazmohamadi 1, Peiman Shahbeigi-Roodposhti-Roodposhti 1
1University of Connecticut

Ceramic-matrix Composite Materials and Their Bending Properties

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Engineering

Focused Ion Beams
Sina Shahbazmohamadi 1, Peiman Shahbeigi-Roodposhti-Roodposhti 1
1University of Connecticut

Focused Ion Beams

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Engineering

Nanocrystalline Alloys and Nano-grain Size Stability
Sina Shahbazmohamadi 1, Peiman Shahbeigi-Roodposhti-Roodposhti 1
1University of Connecticut

Source: Sina Shahbazmohamadi and Peiman Shahbeigi-Roodposhti, School of Engineering, University of Connecticut, Storrs, CT

Alloys with grain size less than 100 nm are known as nanocrystaline alloys. Due to their enhanced physical and mechanical properties, there is an ever-increasing demand to employ them in various industries such as semiconductor, biosensors and aerospace.

To improve the processing and application of nanocrystalline alloys, it is necessary to develop close to 100% dense bulk materials which requires a synergistic effect of elevated temperature and pressure. By increasing the applied temperature and pressure, small grains start to grow and lose their distinguished properties. Thus, it is technologically important to reach a compromise between inter-particle bonding with minimum porosity and loss of nano-scale grain size during consolidating at elevated temperatures.

In this study we aim to eliminate oxygen from solid solution to improve the nano-grain size stability at elevated temperatures. Nano-crystalline Fe-14Cr-4Hf alloy will be synthesized in a protected environment to avoid oxide particles formation.

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Engineering

Directional Solidification and Phase Stabilization
Sina Shahbazmohamadi 1, Peiman Shahbeigi-Roodposhti-Roodposhti 1
1University of Connecticut

Directional Solidification and Phase Stabilization

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Engineering

SEM Imaging of Biological Samples
Peiman Shahbeigi-Roodposhti 1
1University of Connecticut

SEM Imaging of Biological Samples

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Neuroscience

Organotypic Slice Cultures to Study Oligodendrocyte Dynamics and Myelination
Robert A. Hill 1,3, Jelena Medved 1, Kiran D. Patel 1, Akiko Nishiyama 1,2
1Department of Physiology and Neurobiology, University of Connecticut, 2Stem Cell Institute, University of Connecticut, 3Department of Neurology, Yale University School of Medicine

A technique to study NG2 cells and oligodendrocytes using a slice culture system of the forebrain and cerebellum is described. This method allows examination of the dynamics of proliferation and differentiation of cells within the oligodendrocyte lineage where the extracellular environment can be easily manipulated while maintaining tissue cytoarchitecture.

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Neuroscience

3D Modeling of the Lateral Ventricles and Histological Characterization of Periventricular Tissue in Humans and Mouse
Rebecca L. Acabchuk 1, Ye Sun 1, Richard Wolferz, Jr. 1, Matthew B. Eastman 1, Jessica B. Lennington 1, Brett A. Shook 1, Qian Wu 2, Joanne C. Conover 1
1Department of Physiology and Neurobiology, University of Connecticut, 2Department of Anatomic Pathology and Laboratory Medicine, University of Connecticut Health Center

Using MRI scans (human), 3D imaging software, and immunohistological analysis, we document changes to the brain’s lateral ventricles. Longitudinal 3D mapping of lateral ventricle volume changes and characterization of periventricular cellular changes that occur in the human brain due to aging or disease are then modeled in mice.

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Chemistry

Real-time Monitoring of Reactions Performed Using Continuous-flow Processing: The Preparation of 3-Acetylcoumarin as an Example
Trevor A. Hamlin 1, Nicholas E. Leadbeater 1
1Department of Chemistry, University of Connecticut

Real-time monitoring allows for fast optimization of reactions performed using continuous-flow processing. Here the preparation of 3-acetylcoumarin is used as an example. The apparatus for performing in-situ Raman monitoring is described, as are the steps required to optimize the reaction.

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Biology

High-Throughput, Multi-Image Cryohistology of Mineralized Tissues
Nathaniel A. Dyment 1, Xi Jiang 1, Li Chen 1, Seung-Hyun Hong 2, Douglas J. Adams 3, Cheryl Ackert-Bicknell 4, Dong-Guk Shin 2, David W. Rowe 1
1Department of Reconstructive Sciences, University of Connecticut Health Center, 2Department of Computer Science and Engineering, University of Connecticut, 3Department of Orthopaedic Surgery, University of Connecticut Health Center, 4Department of Orthopaedics, University of Rochester

In this manuscript, we present a high-throughput, semi-automated cryohistology platform to produce aligned composite images of multiple response measures from several rounds of fluorescent imaging on frozen sections of mineralized tissues.

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JoVE Core

Magnetic Levitation Coupled with Portable Imaging and Analysis for Disease Diagnostics
Stephanie M. Knowlton 1, Bekir Yenilmez 2, Reza Amin 2, Savas Tasoglu 1,2
1Biomedical Engineering Department, University of Connecticut, 2Mechanical Engineering Department, University of Connecticut

We present a magnetic levitation technique coupled with automated imaging and analysis in both a smartphone-compatible device and a device with embedded imaging and processing. This is applied to measure the density distribution of cells with two demonstrated biomedical applications: sickle cell disease diagnosis and separating white and red blood cells.

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Neuroscience

A Familiarization Protocol Facilitates the Participation of Children with ASD in Electrophysiological Research
Jacqueline Turcios 1,2, Barbara Cook 1, Julia Irwin 2,3, Taylor Rispoli 4, Nicole Landi 2,5
1Department of Communication Sciences and Disorders, Southern Connecticut State University, 2Haskins Laboratories, 3Department of Psychology, Southern Connecticut State University, 4Department of Social Work, Southern Connecticut State University, 5Department of Psychology, University of Connecticut

Electrophysiological research is an important tool for identifying biomarkers of developmental disorders, including Autism Spectrum Disorders (ASD), but data collection in these populations remains challenging. This work presents a familiarization protocol to accompany research that includes electroencephalography (EEG) to improve the likelihood of collecting EEG data from children with ASD.

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Bioengineering

Multimodal Volumetric Retinal Imaging by Oblique Scanning Laser Ophthalmoscopy (oSLO) and Optical Coherence Tomography (OCT)
Weiye Song *1, Libo Zhou *1, Ji Yi 1,2
1Department of Medicine, Boston University School of Medicine, 2Department of Biomedical Engineering, Boston University

Here, we present a protocol to get a large field of view (FOV) three-dimensional (3D) fluorescence and OCT retinal image by using a novel imaging multimodal platform. We will introduce the system setup, the method of alignment, and the operational protocols. In vivo imaging will be demonstrated, and representative results will be provided.

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Genetics

Genetic Engineering of Dictyostelium discoideum Cells Based on Selection and Growth on Bacteria
Peggy Paschke 1, David A. Knecht 2, Thomas D. Williams 1, Peter A. Thomason 3, Robert H. Insall 3, Jonathan R. Chubb 4,5, Robert R. Kay 1, Douwe M. Veltman 1
1MRC Laboratory of Molecular Biology, 2Department of Molecular and Cell Biology, University of Connecticut, 3Cancer Research UK Beatson Institute Glasgow, 4MRC Laboratory for Molecular Cell Biology, University College London, 5Department of Cell and Developmental Biology, University College London

Dictyostelium discoideum is a popular model organism to study complex cellular processes such as cell migration, endocytosis, and development. The utility of the organism is dependent on the feasibility of genetic manipulation. Here, we present methods to transfect Dictyostelium discoideum cells that overcome existing limitations of culturing cells in liquid media.

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Chemistry

Development and Validation of Chromium Getters for Solid Oxide Fuel Cell Power Systems
Ashish Aphale 1, Junsung Hong 1, Boxun Hu 1,2, Prabhakar Singh 1
1Department of Materials Science and Engineering, University of Connecticut, 2Center for Clean Energy Engineering, University of Connecticut

Cathode poisoning from airborne contaminants in trace levels remains a major concern for long-term stability of high-temperature electrochemical systems. We provide a novel method to mitigate the cathode degradations using getters, which capture airborne contaminants at high temperature before entering electrochemically active stack area.

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Neuroscience

Stereological Estimation of Cholinergic Fiber Length in the Nucleus Basalis of Meynert of the Mouse Brain
Prabhakar Singh 1, David W. Peng 1, William Z. Suo 1,2,3,4
1Laboratory for Alzheimer's Disease and Aging Research, Kansas City Veterans Affairs Medical Center, 2Department of Neurology, University of Kansas Medical Center, 3Department of Molecular and Integrative Physiology, University of Kansas Medical Center, 4The University of Kansas Alzheimer's Disease Center

Neuronal fiber length within a three-dimensional structure of a brain region is a reliable parameter to quantify specific neuronal structural integrity or degeneration. This article details a stereological quantification method to measure cholinergic fiber length within the nucleus basalis of Meynert in mice as an example.

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JoVE Core

A Finite Element Approach for Locating the Center of Resistance of Maxillary Teeth
Bill Luu 1, Edward Anthony Cronauer 2, Vaibhav Gandhi 1, Jonathan Kaplan 3, David M. Pierce 3,4, Madhur Upadhyay 1
1Division of Orthodontics, University of Connecticut Health, 2Private Practice, Miami, FL, 3Department of Biomedical Engineering, University of Connecticut, 4Department of Mechanical Engineering, University of Connecticut

This study outlines the necessary tools for utilizing low-dose three-dimensional cone beam-based patient images of the maxilla and maxillary teeth to obtain finite element models. These patient models are then used to accurately locate the CRES of all the maxillary teeth.

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Biology

Using a Thermal Camera to Measure Heat Loss Through Bird Feather Coats
Jordana M.F. Graveley 1, Kevin R. Burgio 1, Margaret Rubega 1
1Department of Ecology and Evolutionary Biology, University of Connecticut

This protocol describes the quantification of heat transmission through a flat skinned avian specimen using a thermal camera and hot water bath. The method allows obtaining of quantitative, comparative data about the thermal performance of feather coats across species using dried flat skin specimens.

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Bioengineering

Fabrication of a Biomimetic Nano-Matrix with Janus Base Nanotubes and Fibronectin for Stem Cell Adhesion
Libo Zhou 1, Anne Yau 1, Wuxia Zhang 1, Yupeng Chen 1
1Department of Biomedical Engineering, University of Connecticut

The goal of this protocol is to show the assembly of a biomimetic nanomatrix (NM) with Janus base nanotubes (JBNTs) and fibronectin (FN). When co-cultured with human mesenchymal stem cells (hMSCs), the NMs exhibit excellent bioactivity in encouraging hMSCs adhesion.

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Biology

Establishing Stable Binary Cultures of Symbiotic Saccharibacteria from the Oral Cavity
Andrew J. Collins *1,2, Pallavi P. Murugkar *1,2, Floyd E. Dewhirst 1,2
1The Forsyth Institute, 2Harvard School of Dental Medicine

We demonstrate a method for isolating difficult-to-grow members of the novel bacterial phylum, Saccharibacteria, by filtering dental plaque and co-culturing with host bacteria.

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Bioengineering

Fabrication and Characterization of Layer-By-Layer Janus Base Nano-Matrix to Promote Cartilage Regeneration
Maxwell Landolina *1, Anne Yau *1, Yupeng Chen 1
1Department of Biomedical Engineering, University of Connecticut

This protocol describes the assembly of a layer-by-layer Janus base nano-matrix (JBNm) scaffold by adding Janus base nanotubes (JBNts), matrilin-3, and Transforming Growth Factor Beta-1 (TGF-β1) sequentially. The JBNm was fabricated and characterized; additionally, it displayed excellent bioactivity, encouraging cell functions such as adhesion, proliferation, and differentiation.

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Neuroscience

Simultaneous Data Collection of fMRI and fNIRS Measurements Using a Whole-Head Optode Array and Short-Distance Channels
Sara Sanchez-Alonso 1, Rebecca R. Canale 2, Isabel F. Nichoson 1, Richard N. Aslin 1,2,3
1Child Study Center, Yale University School of Medicine, 2Department of Psychology, University of Connecticut, 3Department of Psychology, Yale University

We present a method for simultaneously collecting fMRI and fNIRS signals from the same subjects with whole-head fNIRS coverage. The protocol has been tested with three young adults and can be adapted for data collection for developmental studies and clinical populations.

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Neuroscience

Using Live Cell STED Imaging to Visualize Mitochondrial Inner Membrane Ultrastructure in Neuronal Cell Models
Emery L. Ng 1, Ashley L. Reed 2, Christopher B. O’Connell 1, Nathan N. Alder 2
1Center for Open Research Resources and Equipment, University of Connecticut, 2Department of Molecular and Cell Biology, University of Connecticut

This protocol presents a workflow for the propagation, differentiation, and staining of cultured SH-SY5Y cells and primary rat hippocampal neurons for mitochondrial ultrastructure visualization and analysis using stimulated emission depletion (STED) microscopy.

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