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  JoVE Developmental Biology


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 JoVE Biology

Visualization of G3BP Stress Granules Dynamics in Live Primary Cells

1Institut de Génétique Moléculaire de Montpellier, CNRS UMR 5535

JoVE 51197

Stress granules (SGs) are cytoplasmic RNA granules containing stalled ribonucleoprotein particles (RNPs), and important in cellular response to various stresses. Dynamics of SGs can be followed in live cells by visualizing the localization of a tagged component of SGs in transfected primary cells after stress.

 JoVE Biology

Using the Gene Pulser MXcell Electroporation System to Transfect Primary Cells with High Efficiency

1Gene Expression Division, Bio-Rad Laboratories, Inc.

JoVE 1662

This procedure shows how to use the Gene Pulser MXcell electroporation system to rapidly and easily identify the best electroporation conditions for mouse embryonic fibroblasts (MEFs) or other primary cells. Considerations for troubleshooting are also discussed in the associated video.

 JoVE Biology

Viability Assays for Cells in Culture

1Division of Pharmaceutical Sciences, Mylan School of Pharmacy, Duquesne University

JoVE 50645

Therapeutic compounds are often first examined in vitro with viability assays. Blind cell counts by a human observer can be highly sensitive to small changes in cell number but do not assess function. Computerized viability assays, as described here, can assess both structure and function in an objective manner.

 JoVE Biology

Adult and Embryonic Skeletal Muscle Microexplant Culture and Isolation of Skeletal Muscle Stem Cells

1School of Biosciences, University of Birmingham

JoVE 2051

The micro-dissected explants technique is a robust and reliable method for isolating proliferative skeletal muscle cells from juvenile, adult or embryonic muscles as a source of skeletal muscle stem cells. Uniquely, these cells have been clonally derived to produce skeletal muscle stem cell lines used for in vivo transplantation.

 JoVE Medicine

Setting-up an In Vitro Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

1VECT-HORUS SAS, 2Aix-Marseille Université, CNRS, NICN UMR 7259

JoVE 51278

The aim of the present study was to validate the reproducibility of an in vitro BBB model involving a rat syngeneic co-culture of endothelial cells and astrocytes. The endothelial cell monolayer presented high TEER and low LY permeability. Expression of specific TJ proteins, functional responses to inflammation and functionality of transporters and receptors were assessed.

 JoVE Medicine

Isolation and Immortalization of Patient-derived Cell Lines from Muscle Biopsy for Disease Modeling

1Department of Cell Biology, UT Southwestern Medical Center, 2National Institute of Neurological Disorders and Stroke, National Institute of Health, 3Division of Pediatric Pathology, Department of Pathology and Laboratory Medicine, Medical College of Wisconsin, 4Division of Genetics and Genomics, Boston Children's Hospital

JoVE 52307

This protocol describes techniques for live cell isolation and primary culture of myogenic and fibroblast cell lines from muscle or skin tissue. A technique for the immortalization of these cell lines is also described. Altogether, these protocols provide a reliable tool to generate and preserve patient-derived cells for downstream applications.

 JoVE Medicine

Two Methods for Establishing Primary Human Endometrial Stromal Cells from Hysterectomy Specimens

1Department of Pathology, University of Virginia, 2Department of Obstetrics & Gynecology, University of Virginia

JoVE 51513

Establishing primary endometrial stromal cell culture systems from hysterectomy specimens is a valuable biological technique and a crucial step prior to pursuing a vast array of research aims. Here, we describe two methods used to establish stromal cultures from surgically resected endometrial tissues of human patients. 

 JoVE Medicine

An Organotypic High Throughput System for Characterization of Drug Sensitivity of Primary Multiple Myeloma Cells

1Department of Cancer Imaging and Metabolism, H. Lee Moffitt Cancer Center and Research Institute

JoVE 53070

We describe a high-throughput drug sensitivity assay for primary multiple myeloma cells. It consists of a reconstruction of the bone marrow microenvironment (including extracellular matrix and stroma) in multi-well plates, and a non-invasive method for longitudinal quantification of cell viability.

 JoVE Biology

Live Cell Imaging of Primary Rat Neonatal Cardiomyocytes Following Adenoviral and Lentiviral Transduction Using Confocal Spinning Disk Microscopy

1Max-Planck-Institute for Molecular Biomedicine and Institute of Cell Biology, 2Department of Internal Medicine, Yale Cardiovascular Research Center and Section of Cardiovascular Medicine

JoVE 51666

This protocol describes a method of live cell imaging using primary rat neonatal cardiomyocytes following lentiviral and adenoviral transduction using confocal spinning disk microscopy. This enables detailed observations of cellular processes in living cardiomyocytes.

 JoVE Medicine

A Three-dimensional Tissue Culture Model to Study Primary Human Bone Marrow and its Malignancies

1Department of Biological Sciences, Purdue University, 2Department of Oncology, University of Alberta, 3Cross Cancer Institute

JoVE 50947

In standard culture methods cells are taken out of their physiological environment and grown on the plastic surface of a dish. To study the behavior of primary human bone marrow cells we created a 3-D culture system where cells are grown under conditions recapitulating the native microenvironment of the tissue.

 JoVE Bioengineering

Isolation of Primary Human Colon Tumor Cells from Surgical Tissues and Culturing Them Directly on Soft Elastic Substrates for Traction Cytometry

1Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, 2College of Medicine, University of Illinois at Urbana-Champaign, 3Provena Covenant Medical Centre, 4Micro and Nanotechnology Laboratory, University of Illinois at Urbana-Champaign

JoVE 52532

A protocol is described to extract primary human cells from surgical colon tumor and normal tissues. The isolated cells are then cultured on soft elastic substrates (polyacrylamide hydrogels) functionalized by an extracellular matrix protein, and embedded with fluorescent microbeads. Traction cytometry is performed to assess cellular contractile stresses.

 JoVE Biology

Isolation of Myofibroblasts from Mouse and Human Esophagus

1Department of Medicine, Keck School of Medicine, University of Southern California

JoVE 52215

We present a protocol to generate primary cultures of murine and human esophageal stromal cells with a myofibroblast phenotype. Cultured cells have spindle shaped morphology, express α-SMA and vimentin, and lack epithelial, hematopoietic and endothelial cell surface markers. Characterized stromal cells can be used in functional studies of epithelial-stromal interactions.

 JoVE Medicine

Patient Derived Cell Culture and Isolation of CD133+ Putative Cancer Stem Cells from Melanoma

1Institute of Pathology, Laboratory of Molecular Tumor Pathology, Charité - Universitätsmedizin Berlin, 2Institute for Chemistry and Biochemistry, Free University Berlin, 3Laboratory for Functional Genomics Charité (LFGC), Charité - Universitätsmedizin Berlin, 4Comprehensive Cancer Center Charité, Charité - Universitätsmedizin Berlin

JoVE 50200

This article describes the preparation of freshly obtained melanoma tissue into primary cell cultures, and how to remove contaminations of erythrocytes and fibroblasts from the tumor cells. Finally, we describe how CD133+ putative melanoma stem cells are sorted from the CD133- bulk using Magnetic Activated Cell Sorting (MACS).

 JoVE Neuroscience

Modeling Astrocytoma Pathogenesis In Vitro and In Vivo Using Cortical Astrocytes or Neural Stem Cells from Conditional, Genetically Engineered Mice

1Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 2Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, 3Division of Neuropathology, Department of Pathology and Laboratory Medicine, University of North Carolina School of Medicine, 4Curriculum in Genetics and Molecular Biology, University of North Carolina School of Medicine, 5Biological and Biomedical Sciences Program, University of North Carolina School of Medicine, 6Department of Radiation Oncology, Emory University School of Medicine, 7Department of Neurology, Neurosciences Center, University of North Carolina School of Medicine

JoVE 51763

Phenotypically wild-type astrocytes and neural stem cells harvested from mice engineered with floxed, conditional oncogenic alleles and transformed via viral Cre-mediated recombination can be used to model astrocytoma pathogenesis in vitro and in vivo by orthotopic injection of transformed cells into brains of syngeneic, immune-competent littermates.

 JoVE Developmental Biology

Isolation and Quantitative Immunocytochemical Characterization of Primary Myogenic Cells and Fibroblasts from Human Skeletal Muscle

1Centre of Human and Aerospace Physiological Sciences, King's College London, 2Wellcome Trust-Medical Research Council, Cambridge Stem Cell Institute

JoVE 52049

The main adherent cell types derived from human muscle are myogenic cells and fibroblasts. Here, cell populations are enriched using magnetic-activated cell sorting based on the CD56 antigen. Subsequent immunolabelling with specific antibodies and use of image analysis techniques allows quantification of cytoplasmic and nuclear characteristics in individual cells.

 JoVE Immunology and Infection

Preparation and Use of HIV-1 Infected Primary CD4+ T-Cells as Target Cells in Natural Killer Cell Cytotoxic Assays

1Department of Immunology and Microbiology, Rush University Medical Center

JoVE 2668

Cytotoxicity assays to measure natural killer cell lytic responses to HIV-infected cells is limited by the purity of the target cells. We demonstrate here the isolation of a highly purified population of HIV-1 infected primary T-cell blasts by taking advantage of HIV-1 s ability to down-modulate CD4.

 JoVE Bioengineering

The Multi-organ Chip - A Microfluidic Platform for Long-term Multi-tissue Coculture

1Medical Biotechnology, Technische Universität Berlin, 2TissUse GmbH, 3Fraunhofer IWS

JoVE 52526

Here, we present a protocol to coculture primary cells, tissue models and punch biopsies in a microfluidic multi-organ chip for up to 28 days. Human dermal microvascular endothelial cells, liver aggregates and skin biopsies were successfully combined in a common media circulation.

 JoVE Medicine

Method for Obtaining Primary Ovarian Cancer Cells From Solid Specimens

1Department of Obstetrics, Gynecology, and Women's Heath, University of Minnesota, 2Department of Obstetrics and Gynecology, Maricopa Medical Center and St Josephs Hospital and Medical Center, 3Masonic Cancer Center, University of Minnesota

JoVE 51581

This study describes a detailed method for isolation and characterization of primary ovarian cancer cells from solid clinical specimens. Ovarian cancer clinical specimens are subjected to enzymatic digestion to obtain viable, fibroblast-free epithelial ovarian cancer (EOC) cells highly suitable for downstream applications.

 JoVE Bioengineering

Production, Characterization and Potential Uses of a 3D Tissue-engineered Human Esophageal Mucosal Model

1Department of Materials Science and Engineering, University of Sheffield, 2Department of Oncology and Insigneo Institute for in silico Medicine, University of Sheffield, 3Department of Histopathology, Sheffield Teaching Hospitals NHS Foundation Trust

JoVE 52693

This manuscript describes the production, characterization and potential uses of a tissue engineered 3D esophageal construct prepared from normal primary human esophageal fibroblast and squamous epithelial cells seeded within a de-cellularized porcine scaffold. The results demonstrate the formation of a mature stratified epithelium similar to the normal human esophagus.

 JoVE Biology

Primary Human Bronchial Epithelial Cells Grown from Explants

1Medicine, Faculty of Health Sciences, McMaster University

JoVE 1789

Here we describe a detailed method for growing primary human bronchial epithelial cells from explants of human bronchial airway tissue including differentiated growth on an air-liquid interface. This method provides an abundant source of primary cells for investigating the role of the airway epithelium in human lung health and disease.

 JoVE Biology

Strategies for Tracking Anastasis, A Cell Survival Phenomenon that Reverses Apoptosis

1W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins University Bloomberg School of Public Health, 2School of Life Sciences, Chinese University of Hong Kong, 3Center for Cell Dynamics, Department of Biological Chemistry, Johns Hopkins University School of Medicine

JoVE 51964

The term anastasis refers to the phenomenon in which dying cells reverse a cell suicide process at a late stage, repair themselves, and ultimately survive. Here we demonstrate protocols for detecting and tracking cells that undergo anastasis.

 JoVE Biology

Application of Retinoic Acid to Obtain Osteocytes Cultures from Primary Mouse Osteoblasts

1Renal Research Laboratory, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, 2Department of Nephrology, Dialysis and Renal Transplant, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, 3Renal Physiopathology Laboratory, Department of Medical, Surgical and Health Sciences, University of Trieste

JoVE 51465

Treatment of primary mouse osteoblasts with retinoic acid produces a homogeneous population of ramified cells bearing morphological and molecular features of osteocytes. The method overcomes the difficulty of obtaining and maintaining primary osteocytes in culture, and can be advantageous to study cells derived from transgenic models. 

 JoVE Developmental Biology

Zebrafish Keratocyte Explants to Study Collective Cell Migration and Reepithelialization in Cutaneous Wound Healing

1Biomedical Sciences Program, Midwestern University, 2Department of Microbiology & Immunology, Midwestern University

JoVE 52489

Zebrafish keratocytes migrate in cell sheets from explants and provide an in vitro model for the study of the mechanisms of collective cell migration in the context of epithelial wound healing. These protocols detail an effective way to establish primary explant cultures for use in collective cell migration assays.

 JoVE Immunology and Infection

Detection of Fluorescent Nanoparticle Interactions with Primary Immune Cell Subpopulations by Flow Cytometry

1Center for Micro-BioRobotics @SSSA, Istituto Italiano di Tecnologia, 2Department of Biology, University of Pisa, 3Center for Biomolecular Nanotechnologies @UniLe, Istituto Italiano di Tecnologia

JoVE 51345

Analysis of nanoparticle interaction with defined subpopulations of immune cells by flow cytometry.

 JoVE Bioengineering

Improved Method for the Preparation of a Human Cell-based, Contact Model of the Blood-Brain Barrier

1Australian Centre for Blood Diseases, Monash University

JoVE 50934

Establishment of human models of the blood-brain barrier (BBB) can benefit research into brain conditions associated with BBB failure. We describe here an improved technique for preparation of a contact BBB model, which permits coculturing of human astrocytes and brain endothelial cells on the opposite sides of a porous membrane.

 JoVE Immunology and Infection

Flow Cytometric Isolation of Primary Murine Type II Alveolar Epithelial Cells for Functional and Molecular Studies

1Research Group Immune Regulation, Helmholtz Centre for Infection Research, 2Research Group Infection Immunology, Institute of Medical Microbiology, Otto-von-Guericke University, 3Department of Experimental Immunology, Helmholtz Centre for Infection Research

JoVE 4322

We describe the rapid isolation of primary murine type II alveolar epithelial cells (AECII) by flow cytometric negative selection. These AECII show high viability and purity and are suitable for a wide range of functional and molecular studies regarding their role in respiratory conditions such as autoimmune or infectious diseases.

 JoVE Biology

Isolation of Primary Myofibroblasts from Mouse and Human Colon Tissue

1Department of Surgery, UCLA, 2Department of Pathology, UC Irvine

JoVE 50611

The myofibroblast is an influential stromal cell of the gastrointestinal tract that regulates important physiologic processes in both normal and disease states. We describe a technique that allows for the isolation of primary myofibroblasts from both mouse and human colon tissue, which can be utilized for in vitro experimentation.

 JoVE Biology

Rapid Genetic Analysis of Epithelial-Mesenchymal Signaling During Hair Regeneration

1Program in Epithelial Biology, Stanford University School of Medicine

JoVE 4344

Tissue-specific analysis of a hair follicle regeneration model using lentivirus to mediate gain- or loss-of-function.

 JoVE Medicine

Isolation of Human Lymphatic Endothelial Cells by Multi-parameter Fluorescence-activated Cell Sorting

1Murdoch Childrens Research Institute, The Royal Children’s Hospital, 2Department of Paediatrics, Faculty of Medicine, Dentistry and Health Sciences, The University of Melbourne, 3Department of Anatomy and Developmental Biology, Faculty of Medicine, Nursing and Health Sciences, Monash University, Clayton

JoVE 52691

The goal of this protocol is to isolate lymphatic endothelial cells lining human lymphatic malformation cyst-like vessels and foreskins using fluorescence-activated cell sorting (FACS). Subsequent cell culturing and expansion of these cells permits a new level of experimental sophistication for genetic, proteomic, functional and cell differentiation studies.

 JoVE Medicine

Studying Pancreatic Cancer Stem Cell Characteristics for Developing New Treatment Strategies

1Stem Cells & Cancer Group, Molecular Pathology Program, Spanish National Cancer Research Center, 2Institute for Research in Biomedicine (IRB Barcelona), 3Center for Stem Cells in Cancer & Ageing, Barts Cancer Institute, Queen Mary University of London

JoVE 52801

Pancreatic cancer stem cells (CSCs) can be expanded in vitro using the anchorage-independent sphere culture technique, which represents a powerful tool to study CSC biology and can serve as the first step to develop novel CSC-targeting therapies. Here the methodology for expanding, analyzing and targeting of pancreatic CSCs is provided.

 JoVE Biology

Isolation and Culture of Mouse Primary Pancreatic Acinar Cells

1INSERM U1052, Centre de Recherche en Cancérologie de Lyon, 2CNRS UMR5286, Centre de Recherche en Cancérologie de Lyon, 3Université de Lyon, 4Université Lyon 1, 5Centre Léon Bérard

JoVE 50514

In this publication, we describe a rapid and convenient procedure for isolating and culturing primary pancreatic acinar cells from the murine pancreas. This method constitutes a valuable approach to study the physiology of fresh primary normal/untransformed exocrine pancreatic cells.

 JoVE Biology

Culturing Primary Rat Inner Medullary Collecting Duct Cells

1Anchored Signalling, Max-Delbrück-Center for Molecular Medicine, 2Leibniz Institute for Molecular Pharmacology (FMP), 3Charité University Medicine Berlin

JoVE 50366

Arginine-vasopressin (AVP) controls fine-tuning of body water homeostasis through facilitating water reabsorption by renal principal cells. Here, we present a protocol for the cultivation of primary rat inner medullary collecting duct cells suitable for the elucidation of molecular mechanisms underlying AVP-mediated water reabsorption.

 JoVE Biology

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading (TED)

1Institute for Clinical Neurobiology, University of Wuerzburg, 2Department of Synapses - Circuits - Plasticity, Max Planck Institute of Neurobiology, Martinsried, 3Walter Brendel Centre of Experimental Medicine, Ludwig-Maximilians University of Munich

JoVE 50317

Targeted-esterase induced dye loading (TED) supports the analysis of intracellular calcium store dynamics by fluorescence imaging. The method bases on targeting of a recombinant Carboxylesterase to the endoplasmic reticulum (ER), where it improves the local unmasking of synthetic low-affinity Ca2+ indicator dyes in the ER lumen.

 JoVE Medicine

From a 2DE-Gel Spot to Protein Function: Lesson Learned From HS1 in Chronic Lymphocytic Leukemia

1Division of Molecular Oncology, IRCCS, San Raffaele Scientific Institute, 2Department of Haemato-Oncology, King's College London, 3IFOM, FIRC Institute of Molecular Oncology, 4Università Vita-Salute San Raffaele

JoVE 51942

Here we describe a protocol that couples two proteomic techniques, namely 2-dimensional Electrophoresis (2DE) and Mass Spectrometry (MS), to identify differentially expressed/post-translational modified proteins among two or more groups of primary samples. This approach, together with functional experiments, allows the identification and characterization of prognostic markers/therapeutic targets.

 JoVE Bioengineering

Tissue Engineering of a Human 3D in vitro Tumor Test System

1Department of Tissue Engineering and Regenerative Medicine, University Hospital Würzburg

JoVE 50460

Methods to create human 3D tumor tissues as test systems are described. These technologies are based on a decellularized Biological Vascularized Scaffold (BioVaSc), primary human cells and a tumor cell line, which can be cultured under static as well as under dynamic conditions in a flow bioreactor.

 JoVE Biology

Isolation and Culture of Neonatal Mouse Cardiomyocytes

1Randall and Cardiovascular Divisions, King’s College London, 2Division of Cardiology, School of Medicine, University of California San Diego

JoVE 50154

Primary mouse cardiomyocyte cultures are one of the pivotal tools for the investigation of myofibrillar organization and function. The following protocol describes the isolation and culture of primary cardiomyocytes from neonatal mouse hearts. The resulting cardiomyocyte cultures may be subsequently used for a variety of biomechanical, biochemical and cell-biological assays.

 JoVE Neuroscience

Isolation and Culture of Human Fungiform Taste Papillae Cells

1Monell Chemical Senses Center, 2New York University College of Dentistry, 3AFB International

JoVE 3730

We aimed to develop a reproducible protocol for isolating and maintaining long-term cultures of human fungiform taste papillae cells. Cells from human fungiform papillae obtained by biopsy were successfully maintained in culture for more than eight passages (12 months) without loss of viability.

 JoVE Immunology and Infection

An In vitro Model to Study Immune Responses of Human Peripheral Blood Mononuclear Cells to Human Respiratory Syncytial Virus Infection

1Laboratory of Pediatric Infectious Diseases, Department of Pediatrics, Radboud university medical center

JoVE 50766

Human respiratory syncytial virus (HRSV) can cause severe bronchiolitis in young infants. Part of the pathogenesis of severe HRSV disease is caused by the host immune response. Stimulation of primary human immune cells with HRSV provides a fast and reproducible model system to study activation of inflammatory pathways and infection.

 JoVE Biology

Primary Cell Cultures from Drosophila Gastrula Embryos

1Department of Genetics, Harvard Medical School, 2Howard Hughes Medical Institute

JoVE 2215

We provide a detailed protocol for preparing primary cells dissociated from Drosophila embryos. The ability to carry out the effective RNAi perturbation, together with other molecular, biochemical and cell imaging methods will allow a variety of questions to be addressed in Drosophila primary cells.

 JoVE Immunology and Infection

Electroporation of Functional Bacterial Effectors into Mammalian Cells

1Biological Sciences Division, Pacific Northwest National Laboratory, 2Environmental Molecular Science Laboratory, Pacific Northwest National Laboratory, 3Structural Proteomics Group, Ontario Center for Structural Proteomics, University of Toronto, 4Center for Bioproducts and Bioenergy, Washington State University

JoVE 52296

Electroporation was used to insert purified bacterial virulence effector proteins directly into living eukaryotic cells. Protein localization was monitored by confocal immunofluorescence microscopy. This method allows for studies on trafficking, function, and protein-protein interactions using active exogenous proteins, avoiding the need for heterologous expression in eukaryotic cells.

 JoVE Immunology and Infection

An In vitro Model to Study Heterogeneity of Human Macrophage Differentiation and Polarization

1Department of Cardiology, University of Heidelberg

JoVE 50332

Monocyte-derived macrophages are important cells of the innate immune system. Here, we describe an easy to use in vitro model to generate these cells. Using gradient centrifugation, negative bead isolation and specific cell culture conditions, monocyte-derived macrophages can be generated for phenotypic and functional studies.

 JoVE Immunology and Infection

High Yield Purification of Plasmodium falciparum Merozoites For Use in Opsonizing Antibody Assays

1Division of Infection and Immunity, Walter and Eliza Hall Institute of Medical Research, 2Department of Medical Biology, University of Melbourne

JoVE 51590

Measuring antibody function is key to understanding immunity to Plasmodium falciparum malaria. This method describes the purification of viable merozoites, and measurement of opsonization-dependent phagocytosis by flow cytometry.

 JoVE Bioengineering

Generation of a Three-dimensional Full Thickness Skin Equivalent and Automated Wounding

1Department for Tissue Engineering and Regenerative Medicine, University Hospital Würzburg, 2Translational Center Würzburg, Regenerative Therapies in Oncology and Musculoskelettal Disease, Würzburg Branch of the Fraunhofer-Institute Interfacial Engineering and Biotechnology, IGB

JoVE 52576

The goal of this protocol is to build up a three-dimensional full thickness skin equivalent, which resembles natural skin. With a specifically constructed automated wounding device, precise and reproducible wounds can be generated under maintenance of sterility.

 JoVE Immunology and Infection

A Microscopic Phenotypic Assay for the Quantification of Intracellular Mycobacteria Adapted for High-throughput/High-content Screening

1Inserm U1019 - CNRS UMR 8024, Institut Pasteur de Lille, Université de Lille

JoVE 51114

Here, we describe a phenotypic assay applicable to the High-throughput/High-content screens of small-interfering synthetic RNA (siRNA), chemical compound, and Mycobacterium tuberculosis mutant libraries. This method relies on the detection of fluorescently labeled Mycobacterium tuberculosis within fluorescently labeled host cell using automated confocal microscopy.

 JoVE Medicine

In vitro Mesothelial Clearance Assay that Models the Early Steps of Ovarian Cancer Metastasis

1Department of Cell Biology, Harvard Medical School

JoVE 3888

The mesothelial clearance assay described here takes advantage of fluorescently labeled cells and time-lapse video microscopy to visualize and quantitatively measure the interactions of ovarian cancer multicellular spheroids and mesothelial cell monolayers. This assay models the early steps of ovarian cancer metastasis.

 JoVE Biology

Isolation and Differentiation of Stromal Vascular Cells to Beige/Brite Cells

1UCSF Diabetes Center and Department of Cell and Tissue Biology, University of California, San Francisco, 2Department of Biology, University of Copenhagen, Denmark, 3National Institute of Nutrition and Seafood Research, Bergen, Norway

JoVE 50191

Primary white preadipocytes isolated from white adipose tissues in mice can be differentiated into beige/brite cells. Presented here is a reliable cellular model system to study the molecular regulation of "browning" of white fat.

 JoVE Immunology and Infection

Two Methods of Heterokaryon Formation to Discover HCV Restriction Factors

1Division of Experimental Virology, Twincore, Centre for Experimental and Clinical Infection Research, 2Aaron Diamond AIDS Research Center, Laboratory of Retrovirology, The Rockefeller University, NY

JoVE 4029

We describe two methods for conditional trans-complementation of hepatitis C virus (HCV) assembly and the completion of the full viral life cycle, which rely on heterokaryon formation. These techniques are suitable to screen for cell lines that express dominant restriction factors, which preclude production of infectious HCV progeny.

 JoVE Biology

Isolation of Embryonic Ventricular Endothelial Cells

1McAllister Heart Institute, University of North Carolina at Chapel Hill

JoVE 50463

Primary cell culture is a useful technique for analyzing specific populations of cells, particularly from transgenic mouse embryos at specific developmental stages. Herein, embryonic ventricles are dissected and dissociated, and antibody-conjugated beads recognize and separate out the endothelial cells for further analysis.

 JoVE Immunology and Infection

Kupffer Cell Isolation for Nanoparticle Toxicity Testing

1Institute of Pharmaceutical Science, King's College London

JoVE 52989

Liver macrophages, named Kupffer cells, are responsible for the capture of circulating nanoparticles. We describe here a method, of high cell purity and yield, for Kupffer cell isolation. The modified LDH assay is used here to measure the toxicity induced by carbon nanotubes in Kupffer cells.

 JoVE Bioengineering

Gene Transfection toward Spheroid Cells on Micropatterned Culture Plates for Genetically-modified Cell Transplantation

1Graduate School of Medicine, Laboratory of Clinical Biotechnology, The University of Tokyo, 2Graduate School of Engineering, Department of Materials Engineering, The University of Tokyo, 3Graduate School of Engineering, Department of Bioengineering, The University of Tokyo

JoVE 52384

This protocol describes a cell transplantation system using genetically modified, injectable spheroids. Cell spheroids are cultured on micropatterned culture plates and recovered after gene introduction using polyplex nanomicelles. This system facilitates prolonged transgene expression from the transplanted cells in host animals while maintaining the innate function of the cells.

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