1Petroleum & Geosystems Engineering, The University of Texas at Austin
This paper demonstrates the experimental procedure to measure terminal settling velocities of spherical particles in surfactant-based shear thinning viscoelastic fluids. Fluids over a wide range of rheological properties are prepared and settling velocities are measured for a range of particle sizes in unbounded fluids and fluids between parallel walls.
Published January 3, 2014. Keywords: Physics, chemical engineering, settling velocity, Reynolds number, shear thinning, wall retardation
1Fischell Department of Bioengineering, University of Maryland, College Park
Lymph nodes are the immunological tissues that orchestrate immune response and are a critical target for vaccines. Biomaterials have been employed to better target lymph nodes and to control delivery of antigens or adjuvants. This paper describes a technique combining these ideas to inject biocompatible polymer particles into lymph nodes.
Published January 2, 2014. Keywords: Bioengineering, biomaterial, immunology, microparticle, nanoparticle, vaccine, adjuvant, lymph node, targeting, polymer
JoVE Immunology and Infection
Lentiviral expression vectors are the most effective vehicles for stably expressing different effector molecules or reporter constructs in dividing and non-dividing mammalian cells and whole organisms. Here we provide a protocol on how to package lentivector expression constructs in pseudoviral particles and to transduce target cells using the pseudoviral particles.
Published April 8, 2012. Keywords: Immunology, lentivector, virus packaging, pseudovirus production, lentiviral packaging, HIV-based lentivector, lentiviral delivery, lentiviral transduction, lentivirus concentration, stable expression, stable cell lines
JoVE Immunology and Infection
1Laboratory of Virology, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 2Research Technology Branch, Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health
Work with infectious Ebola viruses is restricted to biosafety level 4 laboratories. Tetracistronic minigenome-containing replication and transcription-competent virus like particles (trVLPs) represent a lifecycle modeling system that allows us to safely model multiple infectious cycles under biosafety level 2 conditions, relying exclusively on Ebola virus components.
Published September 27, 2014. Keywords: Infectious Diseases, hemorrhagic Fevers, Viral, Mononegavirales Infections, Ebola virus, filovirus, lifecycle modeling system, minigenome, reverse genetics, virus-like particles, replication, transcription, budding, morphogenesis, entry
1Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, 2Department of Chemistry, The Johns Hopkins University
We describe experimental details of the synthesis of patterned and reconfigurable particles from two dimensional (2D) precursors. This methodology can be used to create particles in a variety of shapes including polyhedra and grasping devices at length scales ranging from the micro to centimeter scale.
Published February 4, 2013. Keywords: Chemistry, Chemical Engineering, Biomolecular Engineering, Materials Science, Physics, Nanotechnology, Molecular Self-assembly, Electrochemistry, Folding, three dimensional, lithography, colloid, patchy particles, particles, nanoparticles, robotics, drug delivery, microfabrication, nanofabrication, nano, assembly, synthesis, reaction, origami
JoVE Immunology and Infection
1Institute of Biochemical Engineering, Technische Universität Braunschweig
A method to precisely generate and to comprehensively characterize morphology of filamentous fungus Aspergillus niger is described, which allows the mathematical correlation of morphological appearance and productivity.
Published March 15, 2012. Keywords: Immunology, morphology engineering, Morphology number (Mn), filamentous fungi, fructofuranosidase, micro particles, image analysis
1Marcoule Institute for Separative Chemistry, UMR 5257 CEA-CNRS-UM2-ENSCM
Acoustic cavitation in liquids submitted to power ultrasound creates transient extreme conditions inside the collapsing bubbles, which are the origin of unusual chemical reactivity and light emission, known as sonoluminescence. In the presence of noble gases, nonequilibrium plasma is formed. The "hot" particles and the photons generated by collapsing bubbles are able to excite species in solution.
Published April 11, 2014. Keywords: Chemistry, Sonochemistry, sonoluminescence, ultrasound, cavitation, nanoparticles, actinides, colloids, nanocolloids
1Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, 2Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign
In this article, we present a microfluidic-based method for particle confinement based on hydrodynamic flow. We demonstrate stable particle trapping at a fluid stagnation point using a feedback control mechanism, thereby enabling confinement and micromanipulation of arbitrary particles in an integrated microdevice.
Published January 21, 2011. Keywords: Bioengineering, hydrodynamic, trap, trapping, confinement, micromanipulation, microfluidics, stagnation point flow
1Department of Chemical Engineering, The Pennsylvania State University
We have used plasma enhanced chemical vapor deposition to deposit thin films ranging from a few nm to several 100 nm on nano-sized particles of various materials. We subsequently etch the core material to produce hollow nanoshells whose permeability is controlled by the thickness of the shell. We characterize the permeability of these coatings to small solutes and demonstrate that these barriers can provide sustained release of the core material over several days.
Published August 16, 2012. Keywords: Physics, Chemical Engineering, Plasma Physics, Plasma coating, Core-shell structure, Hollow particles, Permeability, nanoparticles, nanopowders
1Department of Biochemistry and Molecular Biology, University of British Columbia
Nanodiscs are small discoid particles that incorporate membrane proteins into a small patch of phospholipid bilayer. We provide a visual protocol that shows the step-by-step incorporation of the MalFGK2 transporter into a disc.
Published August 31, 2012. Keywords: Materials science, Nanodiscs, membrane proteins, lipids, ABC transporter, maltose transporter, MalFGK2
JoVE Immunology and Infection
1Plant Sciences Institute, Agricultural Research Service, United States Department of Agriculture, 2Molecular Plant Pathology Laboratory, Agricultural Research Service, United States Department of Agriculture
A method to analyze the solvent accessibility of the thiol group of cysteine residues of Maize rayado fino virus (MRFV)-virus-like particles (VLPs) followed by a peptide cross-linking reaction is described. The method takes advantage of the availability of several chemical groups on the surface of the VLPs that can be targets for specific reactions.
Published February 14, 2013. Keywords: Virology, Plant Biology, Infection, Molecular Biology, Biochemistry, Proteins, Chemicals and Drugs, Analytical, Diagnostic and Therapeutic Techniques and Equipment, Technology, Industry, Agriculture, Chemistry and materials, Virus-like particles (VLPs), VLP, sulfhydryl-reactive chemistries, labeling, cross-linking, multivalent display, Maize rayado fino virus, mosaic virus, virus, nanoparticle, drug delivery, peptides, Nicotiana benthamiana, plant model
1Department of Chemistry, Massachusetts Institute of Technology
Poly(ethylene glycol) (PEG) brush-arm star polymers (BASPs) with narrow mass distributions and tunable nanoscopic sizes are synthesized in via ring opening metathesis polymerization (ROMP) of a PEG-norbornene macromonomer followed by transfer of portions of the resulting living brush initiator to vials containing varied amounts of a rigid, photo-cleavable bis-norbornene crosslinker.
Published October 10, 2013. Keywords: Chemistry, Chemical Engineering, Nanoparticles, Polymers, Drug Delivery Systems, Polymerization, polymers, Biomedical and Dental Materials, brush first, polyethylene glycol, photodegradable, ring opening metathesis polymerization, brush polymer, star polymer, drug delivery, gel permeation chromatography, arm first, core functional, photocleavable
1Department of Nanomedicine, Houston Methodist Research Institute, 2MOE Key Laboratory of Bioinorganic and Synthetic Chemistry, School of Chemistry and Chemical Engineering, Sun Yat-sen University, 3Pediatrics Department of Union Hospital, Huazhong University of Science and Technology, 4CAS Key Laboratory for Biomedical Effects of Nanomaterials & Nanosafety, National Center for Nanoscience & Technology of China, 5Department of Medicine, Weill Cornell Medical College, 6Department of Cell and Developmental Biology, Weill Cornell Medical College
Delivery remains the main challenge for the therapeutic implementation of small interfering RNA (siRNA). This protocol involves the use of a multifunctional and biocompatible siRNA delivery platform, consisting of arginine and polyethylenimine grafted porous silicon microparticles.
Published January 15, 2015. Keywords: Bioengineering, Porous silicon, siRNA, Nanodelivery system, Cancer therapy, Transfection, Polycation functionalization
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.
Published May 20, 2014. Keywords: Chemistry, confocal microscopy, particle tracking, colloids, suspensions, confinement, gelation, microfluidics, image correlation, dynamics, suspension flow
1Research Group Experimental Surgery, Department of Cardiovascular Surgery, Medical Faculty, Heinrich-Heine-University Dusseldorf
A method for isolation of exosomes from whole blood and further analysis by nanoparticle tracking using a semi-automatic instrument is presented in this article. The presented technology provides an extremely sensitive method for visualizing and analyzing particles in liquid suspension.
Published January 17, 2015. Keywords: Biochemistry, Exosome, exosome separation, microvesicle, nanoparticle-tracking analysis, nanoparticle counting, particle size analysis, ultracentrifugation, exosome isolation, blood
1Department of Neurosurgery, University Medical Center Utrecht, 2Brain Center Rudolf Magnus, University Medical Center Utrecht
Extracellular vesicles play important roles in physiological and pathological processes, including coagulation, immune responses, and cancer or as potential therapeutic agents in drug delivery or regenerative medicine. This protocol presents methods for the quantification and size characterization of isolated and non-isolated extracellular vesicles in various fluids using tunable resistive pulse sensing.
Published October 19, 2014. Keywords: Bioengineering, exosomes, microvesicles, extracellular vesicles, quantification, characterization, Tunable Resistive Pulse Sensing, qNano
1Department of Biology, San Diego State University, 2DOE Joint Genome Institute, 3Department of Molecular, Cellular and Developmental Biology, University of Colorado, 4Department of Pediatrics, School of Medicine, University of Colorado
Using the cystic fibrosis airway as an example, the manuscript presents a comprehensive workflow comprising a combination of metagenomic and metatranscriptomic approaches to characterize the microbial and viral communities in animal-associated samples.
Published December 22, 2014. Keywords: Molecular Biology, virome, microbiome, metagenomics, metatranscriptomics, cystic fibrosis, mucosal-surface
1Lawrence Berkeley National Laboratory, The Molecular Foundry
More than half of proteins are small proteins (molecular mass <200 kDa) that are challenging for both electron microscope imaging and three-dimensional reconstructions. Optimized negative staining is a robust and high-throughput protocol to obtain high contrast and relatively high resolution (~1 nm) images of small asymmetric proteins or complexes under different physiological conditions.
Published August 15, 2014. Keywords: Environmental Sciences, small and asymmetric protein structure, electron microscopy, optimized negative staining
JoVE Immunology and Infection
1Department of Pharmacology, University of Minnesota, 2Department of Surgery, University of Minnesota, 33M Corporate Research Laboratory
Here we present a protocol to quantify phagocytosis of fluorescent particles by adherent macrophage cell line using a fluorometric method. This method facilitates a high throughput quantification of particle internalization as well as the resulting actin polymerization.
Published November 27, 2014. Keywords: Immunology, Fluorometry, phagocytosis, high throughput assay, actin polymerization, immunology
1Department of Mechanical Engineering, Vanderbilt University
Combining monodisperse drop generation with inertial ordering of cells and particles, we describe a method to encapsulate a desired number of cells or particles in a single drop at kHz rates. We demonstrate efficiencies twice exceeding those of unordered encapsulation for single- and double-particle drops.
Published June 15, 2012. Keywords: Bioengineering, Drop-based microfluidics, inertial microfluidics, ordering, focusing, cell encapsulation, single-cell biology, cell signaling
1Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles
When working with media and reagents used to culture microorganisms, aseptic technique must be practiced to ensure contamination is minimized. A variety of plating methods are routinely used to isolate, propagate, or enumerate bacteria and phage, all of which incorporate procedures that maintain the sterility of experimental materials.
Published May 11, 2012. Keywords: Basic Protocols, Streak plates, pour plates, soft agar overlays, spread plates, replica plates, bacteria, colonies, phage, plaques, dilutions
1Institut für Geophysik und extraterrestrische Physik, Technische Universität Braunschweig
We present a technique to achieve low-velocity to intermediate-velocity collisions between fragile dust aggregates in the laboratory. For this purpose, two vacuum drop-tower setups have been developed that allow collision velocities between <0.01 and ~10 m/sec. The collision events are recorded by high-speed imaging.
Published June 5, 2014. Keywords: Physics, astrophysics, planet formation, collisions, granular matter, high-speed imaging, microgravity drop tower
1Barrow Brain Tumor Research Center, Barrow Neurological Institute
We describe the production and characterization of nanoparticles and microparticles composed of poly(lactic-co-glycolic acid) using vitamin E-TPGS as an emulsifier. By varying formulation parameters such as the concentration of emulsifier, it is possible to produce nanoparticles with mean diameters ranging from 220 nm to 1.98 µm.
Published December 27, 2013. Keywords: Chemistry, Nanoparticles, Microparticles, PLGA, TPGS, drug delivery, scanning electron microscopy, emulsion, polymers
1Forensic Science Graduate Program, Biochemistry Track, University of Central Florida, 2Department of Chemistry, University of Central Florida, 3National Center for Forensic Science, University of Central Florida
Here we describe an optimized and efficient removal strategy for the collection of bio-particles present in ‘touch DNA’ samples, together with an enhanced amplification protocol involving a one-step 5 µl micro-volume lysis/STR amplification, to permit the recovery of short tandem repeat (STR) profiles of the bio-particle donor(s).
Published March 9, 2015. Keywords: Basic Protocol, Forensic Science, Touch DNA Evidence, Micro-manipulation, Cell Isolation and Recovery, DNA Profiling, Short Tandem Repeat (STR) Analysis
1Institute of Pharmacology and Toxicology and Bio-Imaging Center/Rudolf Virchow Center, DFG-Research Center for Experimental Biomedicine, University of Würzburg, Germany
This protocol describes how to use total internal reflection fluorescence microscopy to visualize and track single receptors on the surface of living cells and thereby analyze receptor lateral mobility, size of receptor complexes as well as to visualize transient receptor-receptor interactions. This protocol can be extended to other membrane proteins.
Published July 25, 2014. Keywords: Bioengineering, pharmacology, microscopy, receptor, live-cell imaging, single-molecule, total internal reflection fluorescence, tracking, dimerization, protein-protein interactions
1Department of Pediatrics, Division of Cardiology, The Children's Hospital of Philadelphia, University of Pennsylvania
These studies report on reversible attachment of adenoviral gene vectors to coatless metal surfaces of stents and model mesh disks. Sustained release of transduction-competent viral particles contingent upon hydrolysis of cross-linkers used for vector immobilization results in a durable site-specific transgene expression in vascular cells and in stented arteries.
Published August 12, 2014. Keywords: Medicine, gene therapy, bioconjugation, adenoviral vectors, stents, local gene delivery, smooth muscle cells, endothelial cells, bioluminescence imaging
1Department of Chemistry and Chemical Engineering, Royal Military College of Canada, 2Analytical Services Unit, School of Environmental Studies, Queen's University
Biochar is a carbon-rich material used as a soil amendment with the ability to sustainably sequester carbon, improve substrate quality and sorb contaminants. This protocol describes the 17 analytical methods used for the characterization of biochar, which is required prior to large scale implementation of these amendments in the environment.
Published November 28, 2014. Keywords: Environmental Sciences, biochar, characterization, carbon sequestration, remediation, International Biochar Initiative (IBI), soil amendment
1Department of Medicine, Brigham and Women's Hospital and Harvard Medical School, 2Laboratory for Biomaterials and Drug Delivery, Department of Anesthesiology, Division of Critical Care, Boston Children's Hospital
The objective of this protocol is to incorporate SDF-1α, a stem cell homing factor, into dextran sulfate-chitosan nanoparticles. The resultant particles are measured for their size and zeta potential, as well as the content, activity, and in vitro release rate of SDF-1α from the nanoparticles.
Published January 22, 2015. Keywords: Chemistry, Dextran sulfate, chitosan, glycan, SDF-1α, nanoparticle, polyelectrolyte complex
1Department of Mechanical Engineering, University of Michigan
A procedure for studying transient flows near boundaries using high-resolution, high-speed particle image velocimetry (PIV) is described here. PIV is a non-intrusive measurement technique applicable to any optically accessible flow by optimizing several parameter constraints such as the image and recording properties, the laser sheet properties, and analysis algorithms.
Published June 24, 2013. Keywords: Physics, Mechanical Engineering, Fluid Mechanics, flow measurement, fluid heat transfer, internal flow in turbomachinery (applications), boundary layer flow (general), flow visualization (instrumentation), laser instruments (design and operation), Boundary layer, micro-PIV, optical laser diagnostics, internal combustion engines, flow, fluids, particle, velocimetry, visualization
1Department of Chemical and Biological Engineering, University of Ottawa, 2Department of Mechanical Engineering, University of Ottawa
Micro-particle image velocimetry (μPIV) is used to visualize paired images of micro particles seeded in blood flows which are cross-correlated to give an accurate velocity profile. Shear rate, maximum velocity, velocity profile shape, and flow rate, each of which has clinical applications, can be derived from these measurements.
Published April 25, 2013. Keywords: Bioengineering, Biophysics, Chemical Engineering, Mechanical Engineering, Biomedical Engineering, Medicine, Anatomy, Physiology, Cellular Biology, Molecular Biology, Hematology, Blood Physiological Phenomena, Hemorheology, Hematocrit, flow characteristics, flow measurement, flow visualization, rheology, Red blood cells, cross correlation, micro blood flows, microfluidics, microhemorheology, microcirculation, velocimetry, visualization, imaging
1Department of Biology, The University of Mississippi
Microplate based procedures are described for the colorimetric or fluorometric analysis of extracellular enzyme activity. These procedures allow for the rapid assay of such activity in large numbers of environmental samples within a manageable time frame.
Published October 1, 2013. Keywords: Environmental Sciences, Environmental Monitoring, Ecological and Environmental Processes, Environmental Microbiology, Ecology, extracellular enzymes, freshwater microbiology, soil microbiology, microbial activity, enzyme activity
1Department of Physiology and Pharmacology, School of Medicine, West Virginia University, 2Center for Cardiovascular and Respiratory Sciences, West Virginia University, 3National Institute for Occupational Safety and Health
A whole-body nanoparticle aerosol inhalation exposure facility was constructed for nano-sized titanium dioxide (TiO2) inhalation toxicology studies. This system provides nano-TiO2 aerosol test atmospheres that have: 1) a steady mass concentration; 2) a homogenous composition free of contaminants; and 3) a stable particle size distribution during aerosol generation.
Published May 7, 2013. Keywords: Medicine, Physiology, Anatomy, Chemistry, Biomedical Engineering, Pharmacology, Titanium dioxide, engineered nanomaterials, nanoparticle, toxicology, inhalation exposure, aerosols, dry powder, animal model
JoVE Immunology and Infection
1Experimental and Clinical Research Center, A joint cooperation between the Charité Medical Faculty and the Max Delbrück Center for Molecular Medicine, 2Berlin Ultrahigh Field Facility (B.U.F.F.), Max Delbrück Center for Molecular Medicine
Tracking of cells using MRI has gained remarkable attention in the past years. This protocol describes the labeling of dendritic cells with fluorine (19F)-rich particles, the in vivo application of these cells, and monitoring the extent of their migration to the draining lymph node with 19F/1H MRI and 19F MRS.
Published March 20, 2013. Keywords: Molecular Biology, Immunology, Cellular Biology, Physiology, Anatomy, Biomedical Engineering, Hematology, nuclear magnetic resonance, NMR, Fluorine, dendritic cells, migration, lymph nodes, magnetic resonance imaging, MRI, magnetic resonance spectroscopy, MRS, spectroscopy, imaging, cell tracking, clinical techniques
1Molecular Biophysics and Biochemistry, Yale University
This article describes a standard method to get a three-dimensional (3D) reconstruction of biological macromolecules using negative staining electron microscopy (EM). In this protocol, we explain how to get the 3D structure of the Saccharomyces cerevisiae exosome complex at medium resolution using the random conical tilt reconstruction method (RCT).
Published March 28, 2011. Keywords: Structural Biology, Electron microscopy, single particle three-dimensional reconstruction, exosome complex, negative staining
1Faculty of Pharmacy, University of Sydney, 2Department of Nanobiomedical Science & BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University
Colloidal probe nanoscopy can be used within a variety of fields to gain insight into the physical stability and coagulation kinetics of colloidal systems and aid in drug discovery and formulation sciences using biological systems. The method described within provides a quantitative and qualitative means to study such systems.
Published July 18, 2014. Keywords: Chemistry, Colloidal Probe, Nanoscopy, Suspension Stability, Adhesion Mapping, Force, Particle Interaction, Particle Kinetics
JoVE Immunology and Infection
1Department of Chemistry, Stony Brook University
Adenovirus particles are engineered to contain either the unnatural amino acid analogue azidohomoalanine or the azido sugar O-GlcNAz. The azide group of each is chemoselectively ligated via "click" chemistry reactions as a means of viral surface modification.
Published August 19, 2012. Keywords: Chemistry, Virology, Immunology, Genetics, adenovirus, azide-alkyne cycloaddition, azido sugar, azidohomoalanine, bioorthogonal, click chemistry, gene therapy, unnatural amino acid
1Department of Biology, San Diego State University, 2Scripps Institution of Oceanography, University of California San Diego
Here, we present a comprehensive protocol to assess the organic and inorganic nutrient availability and the abundance and structure of microbial and viral communities in remote marine environments.
Published November 5, 2014. Keywords: Environmental Sciences, dissolved organic carbon, particulate organic matter, nutrients, DAPI, SYBR, microbial metagenomics, viral metagenomics, marine environment
1Department of Electrical and Computer Engineering, University of Minnesota, 2Department of Biomedical Engineering, University of Minnesota, 3Department of Neurology, Mayo Clinic College of Medicine, 4Department of Immunology, Mayo Clinic College of Medicine
Supported lipid bilayers and natural membrane particles are convenient systems that can approximate the properties of cell membranes and be incorporated in a variety of analytical strategies. Here we demonstrate a method for preparing microarrays composed of supported lipid bilayer-coated SiO2 beads, phospholipid vesicles or natural membrane particles.
Published May 8, 2014. Keywords: Bioengineering, supported lipid bilayer, beads, microarray, fluorescence, microfabrication, nanofabrication, atomic layer deposition, myelin, lipid rafts
1Department of Chemical Engineering, Michigan Technological University, 2Department of Mechanical Engineering, Michigan Technological University, 3XG Sciences, Inc.
A microdevice with high throughput potential is used to demonstrate three-dimensional (3D) dielectrophoresis (DEP) with novel materials. Graphene nanoplatelet paper and double sided tape were alternately stacked; a 700 μm micro-well was drilled transverse to the layers. DEP behavior of polystyrene beads was demonstrated in the micro-well.
Published June 22, 2014. Keywords: Physics, graphene paper, dielectrophoresis, graphene electrodes, 3D laminated microdevice, polystyrene beads, cell diagnostics
1Biomedical Engineering Department, Johns Hopkins University School of Medicine, 2Translational Tissue Engineering Center, Johns Hopkins University School of Medicine, 3Wilmer Eye Institute, Johns Hopkins University School of Medicine, 4Institute for Nanobiotechnology, Johns Hopkins University School of Medicine
A protocol for nanoparticle tracking analysis (NTA) and high-throughput flow cytometry to evaluate polymeric gene delivery nanoparticles is described. NTA is utilized to characterize the nanoparticle particle size distribution and the plasmid per particle distribution. High-throughput flow cytometry enables quantitative transfection efficacy evaluation for a library of gene delivery biomaterials.
Published March 1, 2013. Keywords: Biomedical Engineering, Bioengineering, Tissue Engineering, Cellular Biology, Medicine, Genetics, Biocompatible Materials, Biopolymers, Drug Delivery Systems, Nanotechnology, bioengineering (general), Therapeutics, Nanoparticle, poly(beta-amino ester), high-throughput, transfection, nanoparticle tracking analysis, biomaterial, gene delivery, flow cytometry
1Blood Systems Research Institute, 2Department of Medicine, University of California, San Francisco, 3Department of Laboratory Medicine, University of California, San Francisco
Many different methods exist for the measurement of extracellular vesicles (EVs) using flow cytometry (FCM). Several aspects should be considered when determining the most appropriate method to use. Two protocols for measuring EVs are presented, using either individual detection or a bead-based approach.
Published March 17, 2015. Keywords: Cellular Biology, microvesicles, flow cytometry, exosomes, extracellular vesicles, high throughput, microparticles
1Department of Animal and Avian Sciences, University of Maryland, 2Department of Cell Biology and Molecular Genetics, University of Maryland
C. elegans is usually grown on solid agar plates or in liquid cultures seeded with E. coli. To prevent bacterial byproducts from confounding toxicological and nutritional studies, we utilized an axenic liquid medium, CeHR, to grow and synchronize a large number of worms for a range of downstream applications.
Published August 2, 2014. Keywords: Molecular Biology, C. elegans, axenic media, transgenics, microparticle bombardment, heme, nutrition
1Department of Obstetrics, Perinatal Pharmacology, University Hospital Zurich, 2Laboratory for Materials - Biology Interactions, EMPA Swiss Federal Laboratories for Materials Testing and Research, 3Graduate School for Cellular and Biomedical Sciences, University of Bern
The ex vivo dual recirculating human placental perfusion model can be used to investigate the transfer of xenobiotics and nanoparticles across the human placenta. In this video protocol we describe the equipment and techniques required for a successful execution of a placenta perfusion.
Published June 18, 2013. Keywords: Biomedical Engineering, Medicine, Bioengineering, Anatomy, Physiology, Molecular Biology, Biochemistry, Biophysics, Pharmacology, Obstetrics, Nanotechnology, Placenta, Pharmacokinetics, Nanomedicine, humans, ex vivo perfusion, perfusion, biological barrier, xenobiotics, nanomaterials, clinical model
1School of Biomedical Engineering and Science, Virginia Tech, 2Department of Engineering Science and Mechanics, Virginia Tech
Contactless dielectrophoresis (cDEP) achieves sorting and enrichment of particles via their intrinsic dielectric properties. Fluidic electrode channels replace metallic electrodes traditional to DEP, suiting cDEP to non-damaging sterile characterization and sorting of biological particles. We demonstrate how to prepare a cDEP microdevice and conduct cell characterization and sorting experiments.
Published September 3, 2013. Keywords: Biomedical Engineering, Medicine, Cellular Biology, Molecular Biology, Bioengineering, Anatomy, Physiology, Biophysics, Physics, Microfluidics, Cell Separation, Microfluidic Analytical Techniques, Electrophoresis, Microchip, cancer diagnosis, cell enrichment, cell sorting, microfluidics, dielectrophoresis, Lab on a chip, cells, imaging
1Center for Applied Proteomics and Molecular Medicine, George Mason University, 2Ceres Nanosciences
Several pathological biomarkers cannot be easily detected by current techniques because of their low concentration in biological fluids, the presence of degrading enzymes, and large amounts of high molecular weight proteins. Chemically functionalized hydrogel nanoparticles can harvest, preserve and concentrate low abundance proteins enabling the detection of previously undetectable biomarkers.
Published August 7, 2014. Keywords: Bioengineering, biomarker, hydrogel, low abundance, mass spectrometry, nanoparticle, plasma, protein, urine
1School for Engineering of Matter, Transport and Energy, Arizona State University, 2Department of Neurology, Georgetown University Medical Center, 3Department of Pathology, Georgetown University Medical Center
Using atomic force microscopy in combination with biopanning technology we created a negative and positive biopanning system to acquire antibodies against disease-specific protein variants present in any biological material, even at low concentrations. We were successful in obtaining antibodies to TDP-43 protein variants involved in Amyotrophic Lateral Sclerosis.
Published February 12, 2015. Keywords: Bioengineering, Amyotrophic Lateral Sclerosis, TDP-43, Biopanning, Atomic Force Microscopy, scFv, Neurodegenerative diseases
1Department of Neuroscience and Cell Biology, University of Texas Medical Branch, 2Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, 3Sealy Center for Structural Biology and Molecular Biophysics, University of Texas Medical Branch
We present a combination of Cryo-electron microscopy, lipid nanotechnology, and structure analysis applied to resolve the membrane-bound structure of two highly homologous FVIII forms: human and porcine. The methodology developed in our laboratory to helically organize the two functional recombinant FVIII forms on negatively charged lipid nanotubes (LNT) is described.
Published June 3, 2014. Keywords: Bioengineering, Cryo-electron microscopy, Lipid nanotubes, Helical assembly, Membrane-bound organization, Coagulation factor VIII
1Biomedical Technology, CFD Research Corporation
Flow chambers used in adhesion experiments typically consist of linear flow paths and require multiple experiments at different flow rates to generate a shear adhesion map. SynVivo-SMN enables the generation of shear adhesion map using a single experiment utilizing microliter volumes resulting in significant savings in time and consumables.
Published May 25, 2014. Keywords: Bioengineering, particle, adhesion, shear, microfluidics, vasculature, networks
JoVE Immunology and Infection
1Pneumococcal Research, Murdoch Childrens Research Institute, 2Department of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, The University of Melbourne
Latex agglutination testing is a simple, rapid and inexpensive method for serotyping Streptococcus pneumoniae, and has also been widely applied in diagnostic microbiology. This manuscript describes the in-house production of latex agglutination reagents, quality control procedures and the application of this technique to pneumococcal serotyping.
Published September 25, 2014. Keywords: Immunology, Antisera, pneumococci, polysaccharide capsule, slide agglutination
1Leslie Dan Faculty of Pharmacy, University of Toronto, 2MRC-Laboratory of Molecular Biology, Cambridge, UK
Recent improvements in organotypic brain slice preparations have permitted their exploitation for biotechnological applications. Organotypic slices maintain local structural characteristics of in vivo biology, including functional synaptic connections. Here we present a regioselective biolistic delivery method to label and genetically manipulate these slices.
Published October 24, 2014. Keywords: Neuroscience, Biolistics, gene gun, organotypic brain slices, Diolistic, gene delivery, staining