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Volume:3  Issue:8

April 20, 2013

Cell Biology

Flow Cytometric Detection of Mitochondrial Membrane Potential

Authors: Hsin-Yi Chang
Hsin-Yi ChangAffiliation: Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taipei
Bio-protocol author page: a571
Hsuan-Cheng Huang
Hsuan-Cheng HuangAffiliation: Institute of Biomedical Informatics, National Yang-Ming University, Taipei, Taiwan
Bio-protocol author page: a572
Tsui-Chin Huang
Tsui-Chin HuangAffiliation: Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan
Bio-protocol author page: a573
Pan-Chyr Yang
Pan-Chyr YangAffiliation: Department of Internal Medicine, National Taiwan University Hospital and College of Medicine, Taipei, Taiwan
Bio-protocol author page: a574
Yi-Ching Wang
Yi-Ching WangAffiliation: Department of Internal Medicine, National Taiwan University Hospital and College of Medicine, Taipei, Taiwan
Bio-protocol author page: a575
 and Hsueh-Fen Juan
Hsueh-Fen JuanAffiliation: Department of Life Science, Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan
For correspondence: yukijuan@ntu.edu.tw
Bio-protocol author page: a324
date: 4/20/2013, 10246 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.430 

[Abstract] Mitochondrial membrane potential (Δψm) is an important parameter of mitochondrial function and an indicator of cell health. Depletion of Δψm suggests the loss of mitochondrial membrane integrity reflecting the initiation of the proapoptotic signal. Recently, ...

Article identifier: bio-protocol.org/e430

Flow Cytometric Detection of Reactive Oxygen Species

Authors: Hsin-Yi Chang
Hsin-Yi ChangAffiliation: Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan
Bio-protocol author page: a571
Hsuan-Cheng Huang
Hsuan-Cheng HuangAffiliation: Institute of Biomedical Informatics, National Yang-Ming University, Taipei, Taiwan
Bio-protocol author page: a572
Tsui-Chin Huang
Tsui-Chin HuangAffiliation: Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan
Bio-protocol author page: a573
Pan-Chyr Yang
Pan-Chyr YangAffiliation: Department of Internal Medicine, National Taiwan University Hospital and College of Medicine, Taipei, Taiwan
Bio-protocol author page: a574
Yi-Ching Wang
Yi-Ching WangAffiliation: Department of Pharmacology, National Cheng Kung University, Tainan, Taiwan
Bio-protocol author page: a575
 and Hsueh-Fen Juan
Hsueh-Fen JuanAffiliation: Department of Life Science, Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, Taiwan
For correspondence: yukijuan@ntu.edu.tw
Bio-protocol author page: a324
date: 4/20/2013, 15801 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.431 

[Abstract] Reactive oxygen species (ROS) are molecules containing hydroxyl radicals or peroxides with unpaired electrons. In healthy aerobic cells, ROS are produced naturally as a byproduct of oxidative phosphorylation, oxidoreductase enzymes, or metal catalyzed ...

Article identifier: bio-protocol.org/e431

Immunology

Evaluation of Caspase-1 Activation and IL-1β Production in A Kainic Acid Microdyalisis Brain Injury Model

Authors: Antonio S. Herranz
Antonio S. HerranzAffiliation: Neurobiology Service, Hospital Ramón y Cajal, Madrid, Spain
For correspondence: antonio.sanchez@hrc.es
Bio-protocol author page: a583
Eulalia Bazán
Eulalia BazánAffiliation: Neurobiology Service, Hospital Ramón y Cajal, Madrid, Spain
Bio-protocol author page: a584
Diana Reimers
Diana ReimersAffiliation: Neurobiology Service, Hospital Ramón y Cajal, Madrid, Spain
Bio-protocol author page: a585
María T. Montero-Vega
María T. Montero-VegaAffiliation: Neurobiology Service, Hospital Ramón y Cajal, Madrid, Spain
Bio-protocol author page: a586
Adriano Jménez-Escrig
Adriano Jménez-EscrigAffiliation: Medicina y Especialidades Médicas, Universidad de Alcalá, Madrid, Spain
Bio-protocol author page: a587
 and Pablo Pelegrín
Pablo PelegrínAffiliation: Inflammation and Experimental Surgery Unit, University Clinical Hospital Virgen Arrixaca, Murcia, Spain
For correspondence: pablo.pelegrin@ffis.es
Bio-protocol author page: a326
date: 4/20/2013, 3671 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.433 

[Abstract] Intracerebral infusion of kainic acid (KA) by a microdialysis probe induces a focal swelling in the brain-perfused area which promotes inflammation (Compan et al., 2012; Oprica et al., 2003). The microdialysis technique allows the local in vivo perfusion ...

Article identifier: bio-protocol.org/e433

Microbiology

Plasmodium falciparum Rosette Disruption Assay

Authors: Micheline Guillotte
Micheline GuillotteAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
For correspondence: micheline.guillotte-blisnick@pasteur.fr
Bio-protocol author page: a576
Odile Mercereau-Puijalon
Odile Mercereau-PuijalonAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
Bio-protocol author page: a308
 and Inès Vigan-Womas
Inès Vigan-WomasAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
Bio-protocol author page: a577
date: 4/20/2013, 4196 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.411 

[Abstract] Rosetting, i.e. the capacity of Plasmodium falciparum-infected red blood cells (iRBCs) to bind two or more uninfected red blood cells (RBCs) is associated with severe malaria in African children. Disruption of rosettes using small soluble inhibitors ...

Article identifier: bio-protocol.org/e411

Plasmodium falciparum Rosette Formation Assay

Authors: Inès Vigan-Womas
Inès Vigan-WomasAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
For correspondence: ines.vigan-womas@pasteur.fr
Bio-protocol author page: a577
Micheline Guillotte
Micheline GuillotteAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
Bio-protocol author page: a576
 and Odile Mercereau-Puijalon
Odile Mercereau-PuijalonAffiliation: Parasite Molecular Immunology Unit, Institut Pasteur, Paris, France
Bio-protocol author page: a308
date: 4/20/2013, 5735 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.412 

[Abstract] Rosetting, i.e. the capacity of red blood cells (iRBCs) infected with mature parasite stages to bind two or more uninfected red blood cells (RBCs) is a virulence factor of Plasmodium falciparum. This protocol describes an in vitro assay to monitor rosette ...

Article identifier: bio-protocol.org/e412

Packaging of Retroviral RNA into Viral Particles Analyzed by Quantitative Reverse Transcriptase-PCR

Authors: Bianca Hoffmann
Bianca HoffmannAffiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Bochum, Germany
Bio-protocol author page: a446
 and Bastian Grewe
Bastian GreweAffiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Bochum, Germany
For correspondence: bastian.grewe@rub.de
Bio-protocol author page: a371
date: 4/20/2013, 6282 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.684 

[Abstract] Formation of viral particles and packaging of genomic retroviral RNA into these particles are important steps in the late phase of the viral replication cycle. The efficiency of the incorporation of viral or cellular RNAs into viral particles can be ...

Article identifier: bio-protocol.org/e684

Molecular Biology

High-throughput Method for Determination of Seed Paternity by Microsatellite Markers

Authors: Samik Bhattacharya
Samik BhattacharyaAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
Bio-protocol author page: a582
 and Ian T. Baldwin
Ian T. BaldwinAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
For correspondence: baldwin@ice.mpg.de
Bio-protocol author page: a581
date: 4/20/2013, 5014 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.452 

[Abstract] In this protocol, determination of seed paternity by microsatellite markers in Nicotiana attenuata is described. However, this does not include a protocol for the novel marker selection/identification, but rather exploits the markers generated for a ...

Article identifier: bio-protocol.org/e452

Detection and Cloning of Spliced Transcripts by RT-PCR

Authors: Bianca Hoffmann
Bianca HoffmannAffiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Bochum, Germany
Bio-protocol author page: a446
 and Bastian Grewe
Bastian GreweAffiliation: Department of Molecular and Medical Virology, Ruhr-University Bochum, Bochum, Germany
For correspondence: bastian.grewe@rub.de
Bio-protocol author page: a371
date: 4/20/2013, 4841 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.486 

[Abstract] Using a Reverse Transcriptase-PCR approach spliced transcripts can be converted to cDNA, amplified and cloned into an expression plasmid. Sequencing of the obtained cDNA allows identification of the splicing events that generated the detected RNA (Grewe ...

Article identifier: bio-protocol.org/e486

p65 Chromatin Immunoprecipitation Protocol

Author: Crissy Dudgeon
Crissy DudgeonAffiliation: Pharmacology and Chemical Biology Department, University of Pittsburgh, Pittsburgh, USA
For correspondence: dudgeonc@upmc.edu
Bio-protocol author page: a570
date: 4/20/2013, 3931 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.683 

[Abstract] Chromatin Immunoprecipitation (ChIP) is an important procedure that allows you to verify if a certain protein is physically located at a regulatory region. This information, taken together with other procedures such as luciferase assays and EMSAs, will ...

Article identifier: bio-protocol.org/e683

Neuroscience

Primary Culture of Cortical Neurons

Authors: Rieko Muramatsu
Rieko MuramatsuAffiliation: Department of Molecular Neuroscience, Graduate School of Medicine, Osaka University, Suita, Japan
For correspondence: muramatsu@molneu.med.osaka-u.ac.jp
Bio-protocol author page: a566
 and Toshihide Yamashita
Toshihide YamashitaAffiliation: Department of Molecular Neuroscience, Graduate School of Medicine, Osaka University, Suita, Japan
For correspondence: yamashita@molneu.med.osaka-u.ac.jp
Bio-protocol author page: a379
date: 4/20/2013, 7622 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.496 

[Abstract] Primary culture of neurons from cerebral cortex is a popular model to study neuronal function in vitro and to explore the molecular mechanism of neurite outgrowth in the developing and adult central nervous system. This protocol is for preparing a culture ...

Article identifier: bio-protocol.org/e496

Culture of Rat Olfactory Ensheathing Cells Using EasySep® Magnetic Nanoparticle Separation

Authors: Susan Louise Lindsay
Susan Louise LindsayAffiliation: Glial Cell Biology Group, Institute of Infection, Immunity and Inflammation, University of Glasgow, Glasgow, UK
Bio-protocol author page: a568
 and Susan Carol Barnett
Susan Carol BarnettAffiliation: Glial Cell Biology Group, Institute of Infection, Immunity and Inflammation, University of Glasgow, Glasgow, UK
For correspondence: susan.barnett@glasgow.ac.uk
Bio-protocol author page: a372
date: 4/20/2013, 3339 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.682 

[Abstract] Olfactory ensheathing cells (OECs) can be isolated and purified from a range of postnatal day 7-day to 10-day rat olfactory bulbs. Rat OECs express the CD271/p75NTR receptor and using the “Do-It-Yourself” magnetic nanoparticle EasySep kit from STEMCELL ...

Article identifier: bio-protocol.org/e682

Plant Science

Determination of Nectar Nicotine Concentration in N. attenuata

Authors: Eva Rothe
Eva RotheAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
Bio-protocol author page: a578
Matthias Schöttner
Matthias SchöttnerAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
Bio-protocol author page: a579
Danny Kessler
Danny KesslerAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
Bio-protocol author page: a580
 and Ian T. Baldwin
Ian T. BaldwinAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
For correspondence: baldwin@ice.mpg.de
Bio-protocol author page: a581
date: 4/20/2013, 3997 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.451 

[Abstract] In this protocol, the determination of the nicotine concentration in nectar of Nicotiana attenuata is described. This method is applicable for the investigation of small amounts of nectar (above 1 μl). It is a high-throughput protocol optimized and streamlined ...

Article identifier: bio-protocol.org/e451