Volume: 5 Issue: 21 —BIO-PROTOCOL

Cancer Biology

Isolation and Flow-cytometric Analysis of Mouse Intestinal Crypt Cells Authors: Dayanira Alsina-Beauchamp, Paloma del Reino and Ana Cuenda, date: 11/05/2015, view: 11707, Q&A: 0

The intestinal epithelial layer forms tubular invaginations into the underlying connective tissue of the lamina propria. These structures, termed crypts, are the basic functional unit of the intestine. Colon crypts and the surrounding lamina propria house different cell types, including epithelial cells, stem cells, enterocytes, goblet cells, as ...

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Differentiation of THP1 Cells into Macrophages for Transwell Co-culture Assay with Melanoma Cells Authors: Michael Peter Smith, Helen Young, Adam Hurlstone and Claudia Wellbrock, date: 11/05/2015, view: 32554, Q&A: 2

Understanding how immune cells such as macrophages interact with cancer cells is of increasing interest, as cancer treatments move towards combing both targeted- and immuno-therapies in new treatment regimes. This protocol is using THP-1 cells, a human leukemia monocytic cell line that can be differentiated into macrophages. This allows studying ...

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Capturing the Driving Role of Tumor-host Crosstalk in a Dynamical Model of Tumor Growth Authors: Sebastien Benzekry, Afshin Beheshti, Philip Hahnfeldt and Lynn Hlatky, date: 11/05/2015, view: 6573, Q&A: 0

In 1999, Hahnfeldt et al. proposed a mathematical model for tumor growth as dictated by reciprocal communications between tumor and its associated vasculature, introducing the idea that a tumor is supported by a dynamic, rather than a static, carrying capacity. In this original paper, the carrying capacity was equated with the variable ...

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Cell Biology

Culture of Megakaryocytes from Human Peripheral Blood Mononuclear Cells Authors: Vishal Salunkhe, Petros Papadopoulos and Laura Gutiérrez, date: 11/05/2015, view: 19436, Q&A: 4

Megakaryocytes are the precursor cells of platelets and are bona fide resident cells in the bone marrow but extremely low in numbers (~1% of total nucleated cells). Upon terminal differentiation, megakaryocytes increase their size, become polyploid and develop a demarcation membrane system. Mature megakaryocytes form proplatelets, which ...

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Assessment of Brown Adipocyte Thermogenic Function by High-throughput Respirometry Authors: Kiana Mahdaviani, Ilan Benador and Orian Shirihai, date: 11/05/2015, view: 10619, Q&A: 0

Brown adipose tissue (BAT) has the unique ability to dramatically increase mitochondrial uncoupled fuel oxidation for thermogenesis in response to adrenergic stimulation. A key parameter in assessing brown adipocyte thermogenic capacity is mitochondrial uncoupling as determined by respiration. Measuring mitochondrial oxygen consumption rate (OCR) ...

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Microbiology

Determination of Fructokinase Activity from Zobellia galactanivorans Authors: Agnès Groisillier and Thierry Tonon, date: 11/05/2015, view: 7161, Q&A: 0

Mannitol is a polyol that occurs in a wide range of living organisms, where it fulfills different physiological roles. Several pathways have been described for the metabolism of mannitol by bacteria, including the phosphoenolpyruvate-dependent phosphotransferase system (PST) and a M2DH-based catabolic pathway. The latter involves two enzymes, a ...

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Determination of Mannitol-2-dehydrogenase Activity Authors: Agnès Groisillier and Thierry Tonon, date: 11/05/2015, view: 6401, Q&A: 0

Mannitol is a polyol that occurs in a wide range of living organisms, where it fulfills different physiological roles. Several pathways have been described for the metabolism of mannitol by bacteria, including the phosphoenolpyruvate-dependent phosphotransferase system (PST) and a M2DH-based catabolic pathway. The latter involves two enzymes, a ...

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Substituted Cysteine Accessibility Method for Topology and Activity Studies of Membrane Enzymes Forming Thioester Acyl Intermediates in Bacteria Authors: Sébastien Gélis-Jeanvoine and Nienke Buddelmeijer, date: 11/05/2015, view: 6453, Q&A: 0

The topology of membrane proteins and enzymes can be determined using various methods including reporter protein fusions and accessibility of cysteine residues to alkylating agents. Here we describe a variation of the substituted cysteine accessibility method to determine membrane topology and activity of enzymes containing an active site ...

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Chromatin Immunoprecipitation (ChIP) Assay for Detecting Direct and Indirect Protein – DNA Interactions in Magnaporthe oryzae Authors: Gang Li, Margarita Marroquin-Guzman and Richard A. Wilson, date: 11/05/2015, view: 10665, Q&A: 0

Chromatin immunoprecipitation (ChIP) is a powerful technology for analyzing protein-DNA interactions in cells. Robust ChIP procedures have been established for investigating direct interactions between protein and DNA. However, detecting indirect protein-DNA interactions in vivo is challenging. Recently, we used ChIP to analyze an ...

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Neuroscience

Electroretinogram (ERG) Recordings from Drosophila Authors: Elisheva Rhodes-Mordov, Hadar Samra and Baruch Minke, date: 11/05/2015, view: 11028, Q&A: 0

Phototransduction is a process in which light is converted into electrical signals used by the central nervous system. Invertebrate phototransduction is a process mediated by the phosphoinositide signaling cascade, characterized by Phospholipase C (PLC) as the effector enzyme and the Transient Receptor Potential (TRP) channels as its target. The ...

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Plant Science

Computational Identification of MicroRNA-targeted Nucleotide-binding Site-leucine-rich Repeat Genes in Plants Authors: Zhu-Qing Shao, Yan-Mei Zhang, Bin Wang and Jian-Qun Chen, date: 11/05/2015, view: 7127, Q&A: 0

Plant genomes harbor dozens to hundreds of nucleotide-binding site-leucine-rich repeat (NBS-LRR, NBS for short) type disease resistance genes (Shao et al., 2014; Zhang et al., 2015). Proper regulation of these genes is important for normal growth of plants by reducing unnecessary fitness costs in the absence of pathogen ...

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Observation of Chloroplast Movement in Vallisneria Authors: Yuuki Sakai and Shingo Takagi, date: 11/05/2015, view: 6661, Q&A: 0

Chloroplasts accumulate to weak light and escape from strong light. These light-induced responses have been known from the 19th century (Böhm, 1856). Up to now, many scientists have developed different methods to investigate these dynamic phenomena in a variety of plant species including the model plant Arabidopsis thaliana, a terrestrial ...

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In vitro Phosphorylation Assay of Putative Blue-light Receptor Phototropins Using Microsomal and Plasma-membrane Fractions Prepared from Vallisneria Leaves Authors: Yuuki Sakai, Shin-ichiro Inoue and Shingo Takagi, date: 11/05/2015, view: 6130, Q&A: 0

An aquatic angiosperm Vallisneria (Alismatales: Hydrocharitaceae) has been used as an excellent experimental material over a century to study the light regulation of dynamic intracellular movements including chloroplast redistribution and cytoplasmic streaming (Senn, 1908; Seitz, 1987; Takagi, 1997). However, understanding of the ...

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Stem Cell

Isolation of Murine Adipose Tissue-derived Mesenchymal Stromal Cells (mASCs) and the Analysis of Their Proliferation in vitro Authors: Per Anderson, Ana Belén Carrillo-Gálvez and Francisco Martín, date: 11/05/2015, view: 7800, Q&A: 0

Mesenchymal stromal cells (MSCs) are non-hematopoietic, perivascular cells which support hematopoiesis and are thought to participate in tissue repair in vivo. MSCs can be isolated from various tissues and organs and are defined in vitro as plastic adherent cells expressing CD73, CD90, CD105 (human MSCs) or CD29, CD44, sca-1 ...

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Skeletal Myogenesis in vitro Authors: William C. W. Chen and Bruno Péault, date: 11/05/2015, view: 7213, Q&A: 0

Mature skeletal myofibers are elongated and multinucleated cells. Many stem/progenitor cell types, including committed muscle stem (satellite cells) and progenitor (myoblasts) cells, muscle-derived stem cells, myogenic endothelial cells, and mesenchymal stem/stromal cells, have been shown to exhibit skeletal myogenesis under appropriate inductive ...

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Past Issues in 2015

Volume: 5 Issue: 21

Cancer Biology

Cell Biology

Microbiology

Neuroscience

Plant Science

Stem Cell