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  JoVE Medicine

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  JoVE Developmental Biology


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Electrophoresis, Polyacrylamide Gel: Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.
 JoVE Biology

Denaturing Urea Polyacrylamide Gel Electrophoresis (Urea PAGE)

1School of Biological Sciences, Nanyang Technological University, Singapore - NTU, 2Singapore-MIT Alliance for Reserach and Technology (SMART)

JoVE 1485

Denaturing urea polyacrylamide gel electrophoresis is used to separate single-stranded DNA or RNA up to a limit of 500 nucleotides. Urea in combination with heat denatures samples and unstructured single strands migrate within the gel matrix according to their molecular weight.

 JoVE Biology

Blue Native Polyacrylamide Gel Electrophoresis (BN-PAGE) for Analysis of Multiprotein Complexes from Cellular Lysates

1Spemann Graduate School of Biology and Medicine (SGBM), University of Freiburg, 2Centre for Biological Signalling Studies (bioss) and Biology III, Faculty of Biology, University of Freiburg, 3Department of Molecular Immunology, Max-Planck-Institute of Immunology and Epigenetics

JoVE 2164

In this video, we describe the characterization of multiprotein complexes (MPCs) by blue native polyacrylamide gel electrophoresis (BN-PAGE). In a first dimension, dialyzed cellular lysates are separated by BN-PAGE to identify individual MPCs. In a second dimension SDS-PAGE, MPCs of interest are further subdivided to analyze their constituents by immunoblotting.

 JoVE Immunology and Infection

Nucleocapsid Annealing-Mediated Electrophoresis (NAME) Assay Allows the Rapid Identification of HIV-1 Nucleocapsid Inhibitors

1Department of Pharmaceutical and Pharmacological Sciences, University of Padova, 2The RNA Institute, SUNY Albany

JoVE 52474

This protocol describes NAME, an assay that allows the rapid identification of molecules able to inhibit in vitro the chaperone activities of HIV-1 nucleocapsid protein.

 JoVE Bioengineering

Simple Polyacrylamide-based Multiwell Stiffness Assay for the Study of Stiffness-dependent Cell Responses

1Biomedical Engineering Department, Saint Louis University

JoVE 52643

Here, a method that enables quick, efficient, and inexpensive preparation of polyacrylamide gels in a multiwell plate format is described. The method does not require any specialized equipment and could be easily adopted by any research laboratory. It would be particularly useful in research focused on understanding stiffness-dependent cell responses.

 JoVE Neuroscience

Consensus Brain-derived Protein, Extraction Protocol for the Study of Human and Murine Brain Proteome Using Both 2D-DIGE and Mini 2DE Immunoblotting

1Team Alzheimer & Tauopathies, Jean-Pierre Aubert Research Centre, Inserm UMR 837, 2EA 4308-Department of Reproductive Biology-Spermiology-CECOS, CHRU-Lille, 3EA2686-Laboratorie d'Immunologie, Faculté de Médecine - Pôle Recherche, 4Department of Neurology, CHRU-Lille

JoVE 51339

A common protein extraction protocol using urea/thiourea/SDS buffer for human and mice brain tissue allows indentification of proteins by 2D-DIGE and their subsequent characterization by mini 2DE immunoblotting. This method enables one to obtain more reproducible and reliable results from human biopsies and experimental models.

 JoVE Immunology and Infection

Total Protein Extraction and 2-D Gel Electrophoresis Methods for Burkholderia Species

1Department of Pediatrics, Centre for Understanding and Preventing Infection in Children, University of British Columbia

JoVE 50730

Members of the Burkholderia genus are pathogens of clinical importance. We describe a method for total bacterial protein extraction, using mechanical disruption, and 2-D gel electrophoresis for subsequent proteomic analysis.

 JoVE Biology

Fluorescence Based Primer Extension Technique to Determine Transcriptional Starting Points and Cleavage Sites of RNases In Vivo

1Department of Microbial Genetics, Interfaculty Institute of Microbiology and Infection Medicine Tübingen (IMIT), Faculty of Science, University of Tübingen

JoVE 52134

We here describe a fluorescence based primer extension method to determine transcriptional starting points from bacterial transcripts and RNA processing in vivo using an automated gel sequencer.

 JoVE Application Notes

Simultaneous, Rapid, and Highly Efficient Protein Transfer Using the Trans-Blot Turbo Transfer System - ADVERTISEMENT

1Bio-Rad Laboratories

JoVE 3158

The Trans-Blot Turbo system reduces protein transfer protocols from gels to as little as 3 minutes, while maintaining high efficiency transfers and high throughput. The system enables protein transfer of 2 mini gels in 3 minutes and up to 4 mini gels in as little as 7 minutes.

 JoVE Bioengineering

Porous Silicon Microparticles for Delivery of siRNA Therapeutics

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

JoVE 52075

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.

 JoVE Biology

Electrophoretic Mobility Shift Assay (EMSA) for the Study of RNA-Protein Interactions: The IRE/IRP Example

1Lady Davis Institute for Medical Research, Jewish General Hospital, 2Department of Medicine, McGill University

JoVE 52230

Here we present a protocol to analyze RNA/protein interactions. The electrophoretic mobility shift assay (EMSA) is based on the differential migration of RNA/protein complexes and free RNA during native gel electrophoresis. By using a radiolabeled RNA probe, RNA/protein complexes can be visualized by autoradiography.

 JoVE Chemistry

Steady-state, Pre-steady-state, and Single-turnover Kinetic Measurement for DNA Glycosylase Activity

1Laboratory of Structural Biology, NIEHS, National Institutes of Health

JoVE 50695

Time courses for the glycosylase activity of 8-oxoguanine DNA glycosylase are biphasic exhibiting a burst of product formation and a linear steady-state phase. Utilizing quench-flow techniques, the burst and the steady-state rates can be measured, which correspond to excision of 8-oxoguanine and release of the glycosylase from the product DNA, respectively.

 JoVE Biology

RNA Secondary Structure Prediction Using High-throughput SHAPE

1RT Biochemistry Section, HIV Drug Resistance Program, Frederick National Laboratory for Cancer Research

JoVE 50243

High-throughput selective 2' hydroxyl acylation analyzed by primer extension (SHAPE) utilizes a novel chemical probing technology, reverse transcription, capillary electrophoresis and secondary structure prediction software to determine the structures of RNAs from several hundred to several thousand nucleotides at single nucleotide resolution.

 JoVE Neuroscience

Selection of Aptamers for Amyloid β-Protein, the Causative Agent of Alzheimer's Disease

1Department of Neurology, David Geffen School of Medicine, 2Molecular Biology Institute, University of California, Los Angeles, 3Brain Research Institute, University of California, Los Angeles

JoVE 1955

Aptamers are short ribo-/deoxyribo-oligonucleotides selected by in-vitro evolution methods based on affinity for a specific target. Aptamers are molecular recognition tools with versatile therapeutic, diagnostic, and research applications. We demonstrate methods for selection of aptamers for amyloid β-protein, the causative agent of Alzheimer's disease.

 JoVE Immunology and Infection

Monitoring Activation of the Antiviral Pattern Recognition Receptors RIG-I And PKR By Limited Protease Digestion and Native PAGE

1Institute for Virology, Philipps-University Marburg

JoVE 51415

Innate defenses to virus infections are triggered by pattern recognition receptors (PRRs). The two cytoplasmic PRRs RIG-I and PKR bind to viral signature RNAs, change conformation, oligomerize, and activate antiviral signaling. Methods are described which allow to conveniently monitor the conformational switching and the oligomerization of these cytoplasmic PRRs.

 JoVE Environment

Expression of Recombinant Cellulase Cel5A from Trichoderma reesei in Tobacco Plants

1Institute for Molecular Biotechnology, Bio7, RWTH Aachen University, 2Fraunhofer Institute for Molecular Biology and Applied Ecology

JoVE 51711

Tobacco plants were used to produce a fungal cellulase, TrCel5A, via a transient expression system. The expression could be monitored using a fluorescent fusion protein, and the protein activity was characterized post-expression.

 JoVE Biology

Pouring and Running a Protein Gel by reusing Commercial Cassettes

1Department of Biology, University of Florida, 2UF Genetics Institute, University of Florida, 3Plant Molecular & Cellular Biology Program, University of Florida

JoVE 3465

Our protocol demonstrates how to pour multiple protein gels at a time by recycling Invitrogen Nupage Novex minigel cassettes, and inexpensive materials purchased at a home improvement store. This economical and streamlined method includes a way to store the gels at 4°C for a few weeks. By re-using the plastic gel cassettes from commercially available gels, labs that run frequent protein gels can save significant costs and help the environment.

 JoVE Biology

Studying DNA Looping by Single-Molecule FRET

1School of Physics, Georgia Institute of Technology

JoVE 51667

This study presents a detailed experimental procedure to measure looping dynamics of double-stranded DNA using single-molecule Fluorescence Resonance Energy Transfer (FRET). The protocol also describes how to extract the looping probability density called the J factor.

 JoVE Biology

Fast and Sensitive Colloidal Coomassie G-250 Staining for Proteins in Polyacrylamide Gels

1Biological Medical Research Center (BMFZ), University Duesseldorf

JoVE 1431

This video shall popularize a colloidal Coomassie G-250 staining protocol according to Kang et al. for the detection of average 4 ng protein in gels. The staining is completed within 2 hours and without any effort. We routinely use Kang's protocol for analytical purposes in gel-based proteomics.

 JoVE Immunology and Infection

Proteomic Profiling of Macrophages by 2D Electrophoresis

1Inserm UMR744, University Lille Nord de France

JoVE 52219

Macrophages are the key cells involved in host pathogenicity. Macrophages display phenotypic and functional diversity that can be analysed and detected by proteomic analysis. This article describes how to perform 2D electrophoresis of primary cultures of human macrophages differentiated into M1 or M2 phenotype.

 JoVE Chemistry

Nucleoside Triphosphates - From Synthesis to Biochemical Characterization

1Department of Chemistry and Biochemistry, University of Bern

JoVE 51385

The protocol described herein aims to explain and abridge the numerous obstacles in the way of the intricate route leading to modified nucleoside triphosphates. Consequently, this protocol facilitates both the synthesis of these activated building-blocks and their availability for practical applications.

 JoVE Biology

Preparation of Quality Inositol Pyrophosphates

1Medical Research Council (MRC), Cell Biology Unit and Laboratory for Molecular Cell Biology, University College London

JoVE 3027

Inositol pyrophosphates play an important role in human pathologies such cancer, diabetes and obesity; however, the exact mechanism of action is a matter of dispute. The lack of commercially available inositol pyrophosphates renders detailed studies problematic. Here we describe a simple protocol to produce and isolate milligrams of inositol pyrophosphates.

 JoVE Neuroscience

Telomerase Activity in the Various Regions of Mouse Brain: Non-Radioactive Telomerase Repeat Amplification Protocol (TRAP) Assay

1The Shraga Segal Department of Immunology, Microbiology & Genetics, Faculty of Health Sciences, Ben-Gurion University of the Negev

JoVE 51865

Telomerase is expressed in the neonatal brain and also in distinct regions of the adult brain. We present a non-toxic time saving TRAP assay for the analysis of telomerase activity in various regions of the mouse brain and detection of differences in telomerase activity between male and female mouse brains.

 JoVE Biology

Monitoring Equilibrium Changes in RNA Structure by 'Peroxidative' and 'Oxidative' Hydroxyl Radical Footprinting

1Department of Chemistry, Hunter College, 2Department of Biochemistry, Albert Einstein College of Medicine

JoVE 3244

This protocol describes how to quantify the Mg(II)-dependent formation of RNA tertiary structure by two methods of hydroxyl radical footprinting.

 JoVE Medicine

Skeletal Muscle Gender Dimorphism from Proteomics

1Center for Proteomics, Smith College, 2Department of Molecular Biophysics and Biochemistry, Yale University, 3Department of Chemistry, Smith College, 4Department of Biological Sciences and Center for Proteomics, Smith College

JoVE 3536

A straight-forward set of methods to isolate and determine the identity of the most abundant proteins expressed in skeletal muscle. About 800 spots are discerned on a two-dimensional gel from 10 mg muscle; this allows for the determination of gender-specific protein expression. These methods will give equivalent results in most tissues.

 JoVE Bioengineering

Therapeutic Gene Delivery and Transfection in Human Pancreatic Cancer Cells using Epidermal Growth Factor Receptor-targeted Gelatin Nanoparticles

1Department of Pharmaceutical Sciences, School of Pharmacy, Northeastern University

JoVE 3612

Type B gelatin-based engineered nanovectors system (GENS) was developed for systemic gene delivery and transfection in the treatment of pancreatic cancer. By modification with epidermal growth factor receptor (EGFR) specific peptide on the surface of nanparticles, they could target on EGFR receptor and release plasmid under reducing environment, such as high intracellular glutathione concentrations.

 JoVE Biology

Non-chromatographic Purification of Recombinant Elastin-like Polypeptides and their Fusions with Peptides and Proteins from Escherichia coli

1Department of Biomedical Engineering, Duke University, 2Research Triangle MRSEC, Duke University

JoVE 51583

Elastin-like polypeptides are stimulus-responsive biopolymers with applications ranging from recombinant protein purification to drug delivery. This protocol describes the purification and characterization of elastin-like polypeptides and their peptide or protein fusions from Escherichia coli using their lower critical solution temperature phase transition behavior as a simple alternative to chromatography.

 JoVE Biology

Isolation of Ribosome Bound Nascent Polypeptides in vitro to Identify Translational Pause Sites Along mRNA

1Center for Gene Regulation in Health and Disease, Department of Biological, Geological and Environmental Sciences, Cleveland State University

JoVE 4026

A technique to identify translational pause sites on mRNA is described. This procedure is based on isolation of nascent polypeptides accumulating on ribosomes during in vitro translation of a target mRNA, followed by the size analysis of the nascent chains using a denaturing gel electrophoresis.

 JoVE Biology

Green Fluorescent Protein-based Expression Screening of Membrane Proteins in Escherichia coli

1Oxford Protein Production Facility, Research Complex at Harwell, 2Protein Crystallography Group, Ruhr University, 3MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, 4Membrane Protein Laboratory, Diamond Light Source

JoVE 52357

A streamlined approach to screening for the expression of recombinant membrane proteins in Escherichia coli based on fusion to green fluorescent protein is presented.

 JoVE Biology

Biochemical Reconstitution of Steroid Receptor•Hsp90 Protein Complexes and Reactivation of Ligand Binding

1College of Nursing, Interdisciplinary Life Sciences Research Laboratory, Seattle University, 2College of Science and Engineering, Interdisciplinary Life Sciences Research Laboratory, Seattle University, 3School of Medicine, University of Washington

JoVE 3059

An in vitro method for preparing functional glucocorticoid receptor (GR)•hsp90 protein complexes from purified proteins and cellular lysates is described. The method utilizes immunoadsorption of recombinant GR followed by salt-stripping and protein complex reconstitution. The importance of cofactors and buffer conditions are discussed, as are potential method applications.

 JoVE Bioengineering

Synthesis of an Intein-mediated Artificial Protein Hydrogel

1Artie McFerrin Department of Chemical Engineering, Texas A&M University, College Station, 2Department of Microbial and Molecular Pathogenesis, Texas A&M Health Science Center, Texas A&M University, College Station

JoVE 51202

We present the synthesis of a split-intein-mediated protein hydrogel. The building blocks of this hydrogel are two protein copolymers each containing a subunit of a trimeric protein that serves as a crosslinker and one half of a split intein. Mixing of the two protein copolymers triggers an intein trans-splicing reaction, yielding a polypeptide unit that self-assembles into a hydrogel. This hydrogel is highly pH- and temperature-stable, compatible with organic solvents, and easily incorporates functional globular proteins.

 JoVE Biology

iCLIP - Transcriptome-wide Mapping of Protein-RNA Interactions with Individual Nucleotide Resolution

1Laboratory of Molecular Biology, Medical Research Council - MRC, 2European Bioinformatics Institute, EMBL Heidelberg, 3Computer and Information Science, University of Ljubljana, 4Wellcome Trust Genome Campus, Wellcome Trust Sanger Institute

JoVE 2638

The spatial arrangement of RNA-binding proteins on a transcript is a key determinant of post-transcriptional regulation. Therefore, we developed individual-nucleotide resolution UV crosslinking and immunoprecipitation (iCLIP) that allows precise genome-wide mapping of the binding sites of an RNA-binding protein.

 JoVE Bioengineering

Expression, Isolation, and Purification of Soluble and Insoluble Biotinylated Proteins for Nerve Tissue Regeneration

1Department of Chemical and Biomolecular Engineering, University of Akron

JoVE 51295

Developing biotinylatable fusion proteins has many potential applications in various fields of research. Recombinant protein engineering is a straight forward procedure that is cost-effective, providing high yields of custom-designed proteins.

 JoVE Biology

Protein Membrane Overlay Assay: A Protocol to Test Interaction Between Soluble and Insoluble Proteins in vitro

1Department of Biochemistry and Cell Biology, State University of New York

JoVE 2961

Testing protein-protein interaction is indispensable for dissection of protein functionality. Here, we introduce an in vitro protein-protein binding assay to probe a membrane-immobilized protein with a soluble protein. This assay provides a reliable method to test interaction between an insoluble protein and a protein in solution.

 JoVE Biology

Generation and Purification of Human INO80 Chromatin Remodeling Complexes and Subcomplexes

1Stowers Institute for Medical Research, 2Department of Biochemistry & Molecular Biology, Kansas University Medical Center

JoVE 51720

This protocol describes a procedure for generating and purifying wild type and mutant versions of the human INO80 chromatin remodeling complex. Epitope tagged versions of INO80 subunits are stably expressed in HEK293 cells, and complete complexes and complexes lacking specific sets of subunits are purified by immunoaffinity chromatography.

 JoVE Biology

Expression, Detergent Solubilization, and Purification of a Membrane Transporter, the MexB Multidrug Resistance Protein

1Department of Biological Sciences, University of Illinois Chicago - UIC

JoVE 2134

In this protocol we demonstrate the expression, solubilization, and purification of a recombinantly expressed membrane protein, MexB, as a soluble protein detergent complex. MexB is a multidrug resistance membrane transporter from the opportunistic bacterial pathogen Pseudomonas aeruginosa.

 JoVE Biology

Growth-based Determination and Biochemical Confirmation of Genetic Requirements for Protein Degradation in Saccharomyces cerevisiae

1Department of Biology, Ball State University, 2Division of Nephrology, Cincinnati Children's Hospital

JoVE 52428

This article describes a yeast growth-based assay for the determination of genetic requirements for protein degradation. It also demonstrates a method for rapid extraction of yeast proteins, suitable for western blotting to biochemically confirm degradation requirements. These techniques can be adapted to monitor degradation of a variety of proteins.

 JoVE Immunology and Infection

Purification and Visualization of Lipopolysaccharide from Gram-negative Bacteria by Hot Aqueous-phenol Extraction

1Department of Microbiology, Immunology, & Cancer Biology, University of Virginia Health System

JoVE 3916

We describe a modified hot aqueous-phenol extraction method for purifying lipopolysaccharide (LPS) from Gram-negative bacteria. Once extracted, the LPS can be subsequently analyzed by SDS-PAGE and visualized by direct staining or Western immunoblot.

 JoVE Biology

Assaying Proteasomal Degradation in a Cell-free System in Plants

1Department of Biochemistry and Cell Biology, Stony Brook University, State University of New York

JoVE 51293

Targeted protein degradation represents a major regulatory mechanism for cell function. It occurs via a conserved ubiquitin-proteasome pathway, which attaches polyubiquitin chains to the target protein that then serve as molecular “tags” for the 26S proteasome. Here, we describe a simple and reliable cell-free assay for proteasomal degradation of proteins.

 JoVE Immunology and Infection

Novel Whole-tissue Quantitative Assay of Nitric Oxide Levels in Drosophila Neuroinflammatory Response

1Department of Biological Sciences, University of Alabama

JoVE 50892

Levels of the inflammatory cell-signaling molecule nitric oxide (NO) are commonly assayed using Griess reagent. In this protocol, we have created a modified Griess assay utilizing live Drosophila brain tissue in order to detect the secretion of NO in a simple, quantifiable and highly repeatable method.

 JoVE Biology

Photo-Induced Cross-Linking of Unmodified Proteins (PICUP) Applied to Amyloidogenic Peptides

1Department of Neurology, David Geffen School of Medicine, University of California, Los Angeles, 2Brain Research Institute, Molecular Biology Institute, University of California, Los Angeles, 3Department of Neurology, University of California, Los Angeles

JoVE 1071

Photo-induced cross-linking of unmodified proteins (PICUP) allows characterization of oligomer size distribution in metastable protein mixtures. We demonstrate application of PICUP to three representative amyloidogenic peptides the 40- and 42-residue forms of amyloid β-protein, and calcitonin, and a control peptide growth-hormone releasing factor.

 JoVE Developmental Biology

A Novel Culture Model for Human Pluripotent Stem Cell Propagation on Gelatin in Placenta-conditioned Media

1Department of Biomedical Science, Graduate School of Medicine, Korea University

JoVE 53204

This protocol provides a simple and efficient way to propagate human pluripotent stem cells (hPSCs) using only conditioned media derived from the human placenta in a gelatin-coated dish without additional exogenous supplementation or hPSC-specific synthetic substrata.

 JoVE Biology

Collection and Analysis of Arabidopsis Phloem Exudates Using the EDTA-facilitated Method

1Biochemistry and Molecular Biology, Michigan State Universtiy

JoVE 51111

Knowledge of the composition of the phloem sap as well as the mechanism of its loading and long-distance transport is essential for the understanding of long-distance signaling in plant development and stress/pathogen response and of assimilate transport. This manuscript describes the collection of phloem exudates using the EDTA-facilitated method.

 JoVE Biology

Using SecM Arrest Sequence as a Tool to Isolate Ribosome Bound Polypeptides

1Center for Gene Regulation in Health and Disease, Department of Biological, Geological and Environmental Sciences, Cleveland State University

JoVE 4027

We describe here a technique that is now routinely used to isolate stably bound ribosome nascent chain complexes (RNCs). This technique takes advantage of the discovery that a 17 amino acid long SecM "arrest sequence" can halt translation elongation in a prokaryotic (E. coli) system, when inserted into (or fused to the C-terminus) of virtually any protein.

 JoVE Bioengineering

Fabrication of Biologically Derived Injectable Materials for Myocardial Tissue Engineering

1University of California, San Diego

JoVE 2109

Methods for preparing an injectable matrix gel from decellularized tissue and injecting it into rat myocardium in vivo are described.

 JoVE Neuroscience

SDS-PAGE/Immunoblot Detection of Aβ Multimers in Human Cortical Tissue Homogenates using Antigen-Epitope Retrieval

1Yerkes National Primate Research Center, Emory University, 2Department of Neurology, Institute of Clinical Medicine, Tsukuba University, 3Department of Pathology, New York University School of Medicine, 4Department of Neurology, Emory University

JoVE 1916

We describe a technique for the preparation of clarified human cortical homogenates, protein separation by SDS-PAGE, antigen retrieval and immunoblotting with an antibody to the Aβ peptide. Using this protocol, we consistently detect monomeric and multimeric Aβ in cortical tissue from humans with Alzheimer's pathology.

 JoVE Bioengineering

Low Molecular Weight Protein Enrichment on Mesoporous Silica Thin Films for Biomarker Discovery

1Department of Nanomedicine, The Methodist Hospital Research Institute, 2CAS Key Laboratory for Biological Effects of Nanomaterials & Nanosafety, National Center for Nanoscience and Technology

JoVE 3876

We developed a technology based on mesoporous silica thin film for the selective recovery of low molecular weight proteins and peptides from human serum. The physico-chemical properties of our mesoporous chips were finely tuned to provide substantial control in peptide enrichment and consequently profile the serum proteome for diagnostic purposes.

 JoVE Biology

Demonstration of Proteolytic Activation of the Epithelial Sodium Channel (ENaC) by Combining Current Measurements with Detection of Cleavage Fragments

1Institut für Zelluläre und Molekulare Physiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU)

JoVE 51582

Proteolytic activation of the epithelial sodium channel (ENaC) heterologously expressed in Xenopus laevis oocytes can be demonstrated by combining current measurements with a biotinylation approach to investigate the appearance of ion channel cleavage products at the cell surface. Functionally important cleavage sites can be identified by using site-directed mutagenesis.

 JoVE Biology

Purification and Visualization of Influenza A Viral Ribonucleoprotein Complexes

1Department of Zoology, University of British Columbia - UBC

JoVE 1105

The genome of the influenza A virus consists of eight separate complexes of RNA and proteins, termed viral ribonucleoprotein complexes (vRNPs). This paper describes the glycerol gradient purification and transmission electron microscopy visualization of influenza A vRNPs.

 JoVE Biology

In vitro Methylation Assay to Study Protein Arginine Methylation

1Department of Pulmonary, Critical Care, Sleep, and Allergy, University of Illinois at Chicago, 2Department of Hematology and Oncology, University of Illinois at Chicago, 3Jesse Brown Veterans Affairs Medical Center

JoVE 51997

Protein arginine methylation, catalyzed by a class of enzymes viz., protein arginine methyl transferases (PRMTs), is the process of enzymatic addition of methyl group(s) to arginines within proteins. The in vitro methylation assay is the most dependable tool for assessing the methylation status of known or novel PRMT substrates.

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