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Opening soon: July 2012

JoVE is expanding its scope to include applied physics. We are now accepting technique based manuscripts for our inaugural issues. Open a window into your lab with a visual publication of your protocols.

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Transfecting and Nucleofecting Human Induced Pluripotent Stem Cells


JoVE 3110 10/05/2011

Papri Chatterjee, Yuri Cheung, Chee Liew

UCR Stem Cell Center, Department of Cell Biology and Neuroscience, University of California Riverside

Despite recent advancements in genetic modification, transfection of human embryonic stem cells (HESCs) remains a capricious process. To our knowledge, systematic and efficient methods to transfect human induced pluripotent stem cells (iPSCs) have not been reported. Here, we describe robust protocols to efficiently transfect and nucleofect human iPSCs.

 

Reprogramming Human Somatic Cells into Induced Pluripotent Stem Cells (iPSCs) Using Retroviral Vector with GFP


JoVE 3804 4/03/2012

Kun-Yong Kim, Eriona Hysolli, In-Hyun Park

Yale Stem Cell Center, Department of Genetics, Yale School of Medicine

A method to generate human induced pluripotent stem cells (iPSCs) via retrovirus-mediated ectopic expression of OCT4, SOX2, KLF4 and MYC is described. A practical way to identify human iPSC colonies based on GFP expression is also discussed.

 

Selecting and Isolating Colonies of Human Induced Pluripotent Stem Cells Reprogrammed from Adult Fibroblasts


JoVE 3416 2/20/2012

Urszula Polak1,2, Calley Hirsch1, Sherman Ku3, Joel Gottesfeld3, Sharon Y.R. Dent1, Marek Napierala1

1Department of Molecular Carcinogenesis and Center for Cancer Epigenetics, University of Texas M.D. Anderson Cancer Center, 2Department of Cell Biology, Poznan University of Medical Sciences, 3Department of Molecular Biology, The Scripps Research Institute

We present a protocol for efficient reprogramming of human somatic cells into human induced pluripotent stem cells (hiPSC) using retroviral vectors encoding Oct3/4, Sox2, Klf4 and c-myc (OSKM) and identification of correctly reprogrammed hiPSC by live staining with Tra-1-81 antibody.

 

Feeder-Free Adaptation, Culture and Passaging of Human IPS Cells using Complete KnockOut Serum Replacement Feeder-Free Medium


JoVE 2236 7/15/2010

Kate Wagner, David Welch

GIBCO, Life Technologies

The following protocol provides instruction for adapting human induced Pluripotent Stem (iPS) Cells to feeder-free culture using complete KnockOut Serum Replacement Feeder-Free medium (KSR-FF). Once adapted, instructions for continual maintenance are also provided.

 

Cryopreserving and Recovering of Human iPS Cells using Complete KnockOut Serum Replacement Feeder-Free Medium


JoVE 2237 7/15/2010

Kate Wagner, David Welch

GIBCO, Life Technologies

This protocol describes the detailed procedure for cryopreserving human iPS cells in KnockOut SR cryopreservation medium and recovering these cells in complete KnockOut SR Feeder Free (KSR-FF) medium or feeder-based KnockOut SR medium.

 

Mapping Inhibitory Neuronal Circuits by Laser Scanning Photostimulation


JoVE 3109 10/06/2011

Taruna Ikrar1, Nicholas D. Olivas1, Yulin Shi1, Xiangmin Xu1,2

1Department of Anatomy and Neurobiology, School of Medicine, University of California, Irvine , 2Department of Biomedical Engineering, School of Engineering, University of California, Irvine

This paper introduces an approach of combining laser scanning photostimulation with whole cell recordings in transgenic mice expressing GFP in limited inhibitory neuron populations. The technique allows for extensive mapping and quantitative analysis of local synaptic circuits of specific inhibitory cortical neurons.

 

Teratoma Generation in the Testis Capsule


JoVE 3177 11/07/2011

Suzanne E. Peterson1, Ha T. Tran1, Ibon Garitaonandia1, Sangyoon Han1, Kyle S. Nickey1, Trevor Leonardo2, Louise C. Laurent3, Jeanne F. Loring1

1Department of Chemical Physiology, Scripps Research Institute, 2Department of Chemical Physiology, Scripps Research Institute , 3Department of Reproductive Medicine, University of California

Human pluripotent stem cells (hPSCs) have the potential to treat a myriad of different diseases. The utility of these cells lies in the fact that they can differentiate into any cell type in the body. Here we describe the teratoma assay, which is used to demonstrate the pluripotence of hPSCs.

 

Transduction of Human Cells with Polymer-complexed Ecotropic Lentivirus for Enhanced Biosafety


JoVE 2822 7/24/2011

Bonnie Barrilleaux, Paul Knoepfler

Department of Cell Biology and Human Anatomy, School of Medicine, University of California, Davis

Lentiviruses are a valuable research tool for exploring gene function; however, researchers may wish to avoid production of pantropic lentivirus encoding known or suspected oncogenes. As an alternative, we present a safer protocol for use of ecotropic lentivirus on human cells modified to express the ecotropic receptor mSlc7a1.

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