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Volume:4  Issue:22

November 20, 2014

Cancer Biology

Induction of Colitis and Colitis-associated Colorectal Cancer (CAC)

Authors: Jalaj Gupta
Jalaj GuptaAffiliation: Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain
For correspondence: Gupta@gsh.uni-frankfurt.de
Bio-protocol author page: a1790
 and Angel R. Nebreda
Angel R. NebredaAffiliation 1: Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain
Affiliation 2: Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain
For correspondence: angel.nebreda@irbbarcelona.org
Bio-protocol author page: a1791
date: 11/20/2014, 4975 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1290 

[Abstract] Inflammatory bowel disease (IBD) including Crohn’s disease and ulcerative colitis are characterized by chronic, progressive and relapsing inflammatory disorders. Existing evidence indicate that IBD is associated with a higher risk of developing CAC, ...

Article identifier: bio-protocol.org/e1290

Purification of Tumor-Associated Macrophages (TAM) and Tumor-Associated Dendritic Cells (TADC)

Authors: Damya Laoui
Damya LaouiAffiliation 1: Myeloid Cell Immunology Laboratory, Brussels, Belgium
Affiliation 2: Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
Bio-protocol author page: a1803
Eva Van Overmeire
Eva Van OvermeireAffiliation 1: Myeloid Cell Immunology Laboratory, Brussels, Belgium
Affiliation 2: Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
Bio-protocol author page: a1804
Jiri Keirsse
Jiri KeirsseAffiliation 1: Myeloid Cell Immunology Laboratory, Brussels, Belgium
Affiliation 2: Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
Bio-protocol author page: a1805
Kiavash Movahedi
Kiavash MovahediAffiliation 1: Myeloid Cell Immunology Laboratory, Brussels, Belgium
Affiliation 2: Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
Bio-protocol author page: a1806
 and Jo A Van Ginderachter
Jo A Van GinderachterAffiliation 1: Myeloid Cell Immunology Laboratory, Brussels, Belgium
Affiliation 2: Lab of Cellular and Molecular Immunology, Vrije Universiteit Brussel, Brussels, Belgium
For correspondence: jvangind@vub.ac.be
Bio-protocol author page: a1807
date: 11/20/2014, 7734 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1294 

[Abstract] Tumors are heterogeneous microenvironments where complex interactions take place between neoplastic cells and infiltrating inflammatory cells, such as tumor-associated macrophages (TAM) and tumor-associated dendritic cells (TADC). The relevance of tumor-infiltrating ...

Article identifier: bio-protocol.org/e1294

Cell Biology

Analysis of Intestinal Permeability in Mice

Authors: Jalaj Gupta
Jalaj GuptaAffiliation: Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain
For correspondence: Gupta@gsh.uni-frankfurt.de
Bio-protocol author page: a1790
 and Angel R. Nebreda
Angel R. NebredaAffiliation 1: Institute for Research in Biomedicine (IRB Barcelona), Barcelona, Spain
Affiliation 2: Institució Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain
For correspondence: angel.nebreda@irbbarcelona.org
Bio-protocol author page: a1791
date: 11/20/2014, 12553 views, 3 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1289 

[Abstract] The intestinal epithelial layer serves as a barrier against pathogens and ingested toxins, which are present in the lumen of the intestine. The importance of the intestinal epithelial barrier is emphasized by the alterations in paracellular permeability ...

Article identifier: bio-protocol.org/e1289

Preparation of Golgi Membranes from Rat Liver

Authors: Carmen Valente
Carmen ValenteAffiliation: Institute of Protein Biochemistry (CNR), Naples, Italy
Bio-protocol author page: a1792
Gabriele Turacchio
Gabriele TuracchioAffiliation: Institute of Protein Biochemistry (CNR), Naples, Italy
Bio-protocol author page: a1793
Stefania Spanò
Stefania SpanòAffiliation: Institute of Protein Biochemistry (CNR), Naples, Italy
Bio-protocol author page: a1794
Alberto Luini
Alberto LuiniAffiliation: Institute of Protein Biochemistry (CNR), Naples, Italy
Bio-protocol author page: a1795
 and Daniela Corda
Daniela CordaAffiliation: Institute of Protein Biochemistry (CNR), Naples, Italy
For correspondence: d.corda@ibp.cnr.it
Bio-protocol author page: a1796
date: 11/20/2014, 2391 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1291 

[Abstract] This protocol details the isolation of enriched Golgi membranes from rat liver, using discontinuous density gradient centrifugation. This high-yield extraction method is useful for several applications, including immunoprecipitation of solubilised Golgi ...

Article identifier: bio-protocol.org/e1291

Separation of the Inner and Outer Mitochondrial Membrane in HeLa Cells

Authors: Naotaka Nishimura
Naotaka NishimuraAffiliation: Department of Molecular Genetics, Kumamoto University, Kumamoto, Japan
Bio-protocol author page: a1816
 and Masato Yano
Masato YanoAffiliation: Department of Medical Technology, Kumamoto Health Science University, Kumamoto, Japan
For correspondence: yano@kumamoto-hsu.ac.jp
Bio-protocol author page: a1817
date: 11/20/2014, 7607 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1299 

[Abstract] Mitochondria are organelles that have important functions in oxidative phosphorylation, fatty acid oxidation and apoptosis signaling. They have two distinct membranes, outer membrane (OM) and inner membrane (IM). IM contains respiratory chain complexes ...

Article identifier: bio-protocol.org/e1299

Assay of Ornithine Decarboxylase and Spermidine/Spermine N1-acetyltransferase Activities

Authors: Mervi T. Hyvönen
Mervi T. HyvönenAffiliation: School of Pharmacy, University of Eastern Finland, Kuopio, Finland
For correspondence: mervi.hyvonen@uef.fi
Bio-protocol author page: a1820
Tuomo Keinänen
Tuomo KeinänenAffiliation: School of Pharmacy, University of Eastern Finland, Kuopio, Finland
Bio-protocol author page: a1821
 and Leena Alhonen
Leena AlhonenAffiliation: School of Pharmacy, University of Eastern Finland, Kuopio, Finland
Bio-protocol author page: a1822
date: 11/20/2014, 3082 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1301 

[Abstract] The polyamines, spermidine (Spd) and spermine, and their diamine precursor putrescine, are important regulators of various cellular functions, such as proliferation and differentiation. Polyamine homeostasis is tightly regulated on the level of uptake, ...

Article identifier: bio-protocol.org/e1301

Microbiology

Purification and Sequencing of DNA Guides from Prokaryotic Argonaute

Authors: Daan C. Swarts
Daan C. SwartsAffiliation: Department of Agrotechnology and Food Sciences, Wageningen University, Wageningen, the Netherlands
For correspondence: daan.swarts@wur.nl
Bio-protocol author page: a1688
Edze R. Westra
Edze R. WestraAffiliation: Department of Agrotechnology and Food Sciences, Wageningen University, Wageningen, the Netherlands
Bio-protocol author page: a1801
Stan J. J. Brouns
Stan J. J. BrounsAffiliation: Department of Agrotechnology and Food Sciences, Wageningen University, Wageningen, the Netherlands
Bio-protocol author page: a1802
 and John van der Oost
John van der OostAffiliation: Department of Agrotechnology and Food Sciences, Wageningen University, Wageningen, the Netherlands
Bio-protocol author page: a1690
date: 11/20/2014, 3429 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1293 

[Abstract] Some proteins utilize nucleic acids to guide them to complementary nucleic acid targets. One example is prokaryotic Argonaute protein, which, binds small single stranded DNA molecules as guides (Swarts et al., 2014). This protocol describes a method ...

Article identifier: bio-protocol.org/e1293

Infectious Focus Assays and Multiplicity of Infection (MOI) Calculations for Alpha-herpesviruses

Authors: Anna Sloutskin
Anna SloutskinAffiliation: Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel
Bio-protocol author page: a1808
 and Ronald S. Goldstein
Ronald S. GoldsteinAffiliation: Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel
For correspondence: ron.goldstein@biu.ac.il
Bio-protocol author page: a1809
date: 11/20/2014, 11608 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1295 

[Abstract] Titration of viral stocks is a critical process before any experimental use of the virus. Here we describe an infectious focus assay for several alphaherpesviruses, a titration method for fluorescently labeled viruses, based on the original plaque assay. ...

Article identifier: bio-protocol.org/e1295

Semi-denaturing Detergent Agarose Gel Electrophoresis (SDD-AGE)

Authors: Laura Molina-García
Laura Molina-GarcíaAffiliation: Department of Cellular and Molecular Biology, Centro de Investigaciones Biológicas-CSIC, Madrid, Spain
Bio-protocol author page: a1812
 and Fátima Gasset-Rosa
Fátima Gasset-RosaAffiliation: Department of Neurosciences, University of California, San Diego, USA
For correspondence: fgasset@ucsd.edu
Bio-protocol author page: a1813
date: 11/20/2014, 4511 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1297 

[Abstract] Pathological proteins in neurodegenerative diseases suffer a conformational change to a misfolded amyloid state. Such pathological event leads to the aggregation of these proteins that indefinitely propagates as an altered form of itself, and harbor ...

Article identifier: bio-protocol.org/e1297

Competitive ELISA for Protein-lipopolysaccharide (LPS) Binding

Authors: Victoria Martínez-Sernández
Victoria Martínez-SernándezAffiliation: Departamento de Microbiología y Parasitología, Universidad de Santiago de Compostela, Santiago de Compostela, Spain
Bio-protocol author page: a1814
 and Florencio M. Ubeira
Florencio M. UbeiraAffiliation: Departamento de Microbiología y Parasitología, Universidad de Santiago de Compostela, Santiago de Compostela, Spain
For correspondence: fm.ubeira@usc.es
Bio-protocol author page: a1815
date: 11/20/2014, 3655 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1298 

[Abstract] Lipopolysaccharide is the major constituent of the outer membrane of gram-negative bacteria and, once released from the bacterial surface into the bloodstream, is a potent activator of the host immune system, which can lead to septic shock. LPS has a ...

Article identifier: bio-protocol.org/e1298

Isolation and Characterisation of Dendritic Cells from Peripheral Blood

Authors: Matthew Reeves
Matthew ReevesAffiliation: Institute of Immunity and Transplantation, University College London, London, UK
For correspondence: matthew.reeves@ucl.ac.uk
Bio-protocol author page: a1818
 and John Sinclair
John SinclairAffiliation: Institute of Immunity and Transplantation, University College London, London, UK
Bio-protocol author page: a1819
date: 11/20/2014, 4890 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1300 

[Abstract] Latency and reactivation of human cytomegalovirus (HCMV) is intimately associated with the myeloid lineage. Multiple studies have used in vitro protocols to generate dendritic cells (DCs) from myeloid precursors. Here we describe the direct isolation ...

Article identifier: bio-protocol.org/e1300

Secretion Assay in Shigella flexneri

Authors: Jonathan Reinhardt
Jonathan ReinhardtAffiliation: Max-Planck-Institute for Infection Biology, Berlin, Germany
Bio-protocol author page: a1823
 and Michael Kolbe
Michael KolbeAffiliation: Max-Planck-Institute for Infection Biology, Berlin, Germany
For correspondence: kolbe@mpiib-berlin.mpg.de
Bio-protocol author page: a1824
date: 11/20/2014, 3200 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1302 

[Abstract] Shigella flexneri (S. flexneri) is a Gram-negative bacterium that causes gastroenteritis and shigellosis in humans. In order to establish and maintain an infection, S. flexneri utilises a type three secretion system (T3SS) to deliver virulence factors ...

Article identifier: bio-protocol.org/e1302

Plant Science

In vitro Detection of S-acylation on Recombinant Proteins via the Biotin-Switch Technique

Authors: Dong Qi
Dong QiAffiliation: Department of Biology, Indiana University, Bloomington, USA
Present address: Eurofins Lancaster Laboratory, Inc., Altria Client Services, Richmond, Virginia
Bio-protocol author page: a1810
 and Roger W. Innes
Roger W. InnesAffiliation: Department of Biology, Indiana University, Bloomington, USA
For correspondence: rinnes@indiana.edu
Bio-protocol author page: a1811
date: 11/20/2014, 2707 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1296 

[Abstract] Protein palmitoylation is the post-translational modification of proteins via the attachment of palmitate through acyl linkages. The nucleophile sulfhydryl group of cysteines is the common palmitoylation site. Covalent attachment of palmitate occurs ...

Article identifier: bio-protocol.org/e1296

Stem Cell

Isolation of FAP Cells from Mouse Dystrophic Skeletal Muscle Using Fluorescence Activated Cell Sorting

Authors: Nicoletta Cordani
Nicoletta CordaniAffiliation 1: Scientific Institute, Lecco, Italy
Affiliation 2: Department of Health of Science, University of Milano-Bicocca, Monza, Italy
Bio-protocol author page: a1797
Viviana Pisa
Viviana PisaAffiliation: Department of Biomedical and Clinical Sciences, University of Milano, Milan, Italy
Bio-protocol author page: a1798
Laura Pozzi
Laura PozziAffiliation: Scientific Institute, Lecco, Italy
Bio-protocol author page: a1799
 and Clara Sciorati
Clara ScioratiAffiliation: Division of Regenerative Medicine, Ospedale San Raffaele, Milan, Italy
For correspondence: sciorati.clara@hsr.it
Bio-protocol author page: a1800
date: 11/20/2014, 3160 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1292 

[Abstract] A population of muscle resident CD45-, CD31- cells expressing the mesenchymal PDGF receptor alpha (PDGFRα) as well as Sca-1 was first isolated in healthy mouse muscles in Uezumi et al. (2010). In the same year, Joe et al. (2010) identified and purified ...

Article identifier: bio-protocol.org/e1292