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Cancer Biology

Longitudinal Bioluminescent Quantification of Three Dimensional Cell Growth Authors:  Michael K. Wendt and William P. Schiemann, date: 12/05/2013, view: 4482, Q&A: 0
The use of three-dimensional (3D) cell culture systems is widely accepted as representing a more physiologically relevant means to propagate mammary epithelial and breast cancer cells. However, 3D cultures systems are plagued by several experimental and technical limitations as compared to their traditional 2D counterparts. For instance, ...
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Cell Biology

ATP and Lactate Quantification Authors:  Isaline Rowe, Marco Chiaravalli and Alessandra Boletta, date: 12/05/2013, view: 8599, Q&A: 0
Cells use glucose to generate energy by two different metabolic processes: lactic fermentation and aerobic respiration. In the first common series of reactions, glucose is converted into pyruvate. In anaerobic conditions, pyruvate is transformed into lactate, this process yields to 2 ATP molecules per glucose molecule. In the presence of oxygen, ...
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Mitochondrial Transmembrane Potential (ψm) Assay Using TMRM Authors:  Isaline Rowe and Alessandra Boletta, date: 12/05/2013, view: 14093, Q&A: 0
During cellular respiration, nutrients are oxidized to generate energy through a mechanism called oxidative phosphorylation, which occurs in the mitochondria. During oxidative phosphorylation, the gradual degradation of molecules through the TCA cycle releases electrons from the covalent bonds that are broken. These electrons are captured by NAD+ ...
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Isolation of CD34+ Cells from Human Fetal Liver and Cord Blood Authors:  Qingfeng Chen and Jianzhu Chen, date: 12/05/2013, view: 6585, Q&A: 2
CD34 is a glycosylated cell surface protein and represents a well-known marker for primitive progenitor cells in various organs, especially cord blood, bone marrow and fetal liver. CD34+ progenitor cells are suitable for a series of studies, e.g. cell differentiation, transplantation as well as construction of humanized mouse ...
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Developmental Biology

Acquisition of Leftward Flow in Xenopus laevis Authors:  Thomas Thumberger and Martin Blum, date: 12/05/2013, view: 4517, Q&A: 0
In Xenopus, the left-right axis is established following an extracellular vectorial leftward flow driven by monocilia at the gastrocoel roof plate (GRP) during late gastrulation / neurulation (Schweickert et al., 2007). As the GRP lies inside the developing archenteron, imaging of flow is challenging. Here we present the detailed ...
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Immunology

Purification of Human Monocytes and Lymphocyte Populations by Counter Current Elutriation – A Short Protocol Authors:  Elizabeth V. Clarke, Marie E. Benoit and Andrea J. Tenner, date: 12/05/2013, view: 5691, Q&A: 0
Investigations of the activation processes involved in human monocytes and monocyte-derived macrophages and dendritic cells often required large numbers of cells that have not been possibly altered or activated by adherence to surfaces, by binding of antibodies to surface antigens during positive selection, or by release of activators by platelets ...
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Chitinase Assay from Cultured Bone Marrow Derived Macrophages Authors:  Danielle Worth, J. Philip Nance and Emma H. Wilson, date: 12/05/2013, view: 3835, Q&A: 0
Chitinases are chitin-degrading enzymes. Chitinases play essential roles in combating chitin-containing pathogens as well as established roles in asthmatic inflammation. This assay is designed to detect chitinase activity in macrophage cell lysates. The chitin substrate is labeled with 4-methylumbelliferone. Hydrolysis of chitin releases ...
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Determination of the Intracellular Calcium Concentration in Peritoneal Macrophages Using Microfluorimetry Authors:  Silvia González-Ramos, Luz María G. Carrasquero, Esmerilda G. Delicado, María T. Miras-Portugal, María Fernández-Velasco and Lisardo Boscá, date: 12/05/2013, view: 8393, Q&A: 1
Calcium is one of the most important intracellular messengers in biological systems. Ca2+ microfluorimetry is a valuable tool to assess information about mechanisms involved in the regulation of intracellular Ca2+ levels in research on cells and in living tissues. In essence, the use of a dye that fluoresces in the presence ...
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Escherichia coli Outer Membrane Vesicle Immunization Protocol and Induction of Bacterial Sepsis Authors:  Oh Youn Kim, Bok Sil Hong, Kyong-Su Park, Yae Jin Yoon, Seng Jin Choi, Won Hee Lee, Tae-Young Roh, Yoon-Keun Kim and Yong Song Gho, date: 12/05/2013, view: 6307, Q&A: 0
Outer membrane vesicles (OMVs) are spherical bilayered phospholipids of 20-200 nm in size produced from all Gram-negative bacteria and Gram-positive bacteria investigated to date. OMVs, which resemble the outer membrane and periplasm in composition, are proinflammatory and immunogenic facsimiles, and therefore could activate both innate and ...
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Microbiology

Adherent-invasive Escherichia coli Biofilm Formation Assays Authors:  Benoit Chassaing and Arlette Darfeuille-Michaud, date: 12/05/2013, view: 6458, Q&A: 0
Patients with Crohn’s disease are abnormally colonized by adherent-invasive Escherichia coli (AIEC) bacteria (Chassaing and Darfeuille-Michaud, 2011). These bacteria are able to adhere to and invade intestinal epithelial cells (IEC), to replicate within macrophages, and were recently described to be able to form biofilms (Martinez-Medina ...
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Burkholderia glumae Competent Cells Preparation and Transformation Authors:  Hiromasa Saitoh and Ryohei Terauchi, date: 12/05/2013, view: 5389, Q&A: 1
Bukholderia glumae is a gram-negative bacterium which causes grain rot, seedling rot and panicle blight in rice and bacterial wilt in many field crops. This bacterium has been reported from major rice growing regions around the world and is now considered as an emerging major pathogen of rice (Tsushima et al., 1996; Jeong et al. ...
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Preparation of Outer Membrane Vesicle from Escherichia coli Authors:  Oh Youn Kim, Bok Sil Hong, Kyong-Su Park, Yae Jin Yoon, Seng Jin Choi, Won Hee Lee, Tae-Young Roh, Yoon-Keun Kim and Yong Song Gho, date: 12/05/2013, view: 9557, Q&A: 0
Outer membrane vesicles (OMVs) are spherical bilayered phospholipids of 20-200 nm in size produced from all Gram-negative bacteria and Gram-positive bacteria investigated to date. Previous biochemical and proteomic studies have revealed that the Gram-negative bacteria-derived OMVs are composed of various components like outer membrane proteins, ...
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Neuroscience

Preparation of Primary Neurons from Rat Median Preoptic Nucleus (MnPO) Author:  Emmanuelle Berret, date: 12/05/2013, view: 5047, Q&A: 0
Studying cell physiology is really important to understand their function and to determine action mechanisms taking place in these cells. Using brain slices can be sometime difficult to directly access to cells like neurons. Even if it more steps are needed to get cultured cells, this type of preparation allow a better access to neurons and ...
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Autoradiographic 3H-Gaboxadol Receptor Binding Protocol Authors:  Lynne Ling and Donald Caspary, date: 12/05/2013, view: 3633, Q&A: 0
Gaboxadol (4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol, THIP), a GABAA receptor δ-subunit specific agonist, when present at low (μM) concentrations, preferentially binds and activates extrasynaptic (non-γ2, δ-subunit-containing) GABAARs (Storustovu and Ebert, 2006; Richardson et al., 2011, 2013).

In ...
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Plant Science

Measurement of Potassium Content in Arabidopsis Authors:  Mitsuru Abo, Yuriko Osakabe, Kazuko Y-Shinozaki and Etsuro Yoshimura, date: 12/05/2013, view: 5352, Q&A: 0
Potassium is an essential element in plant growth and has an important role in regulating cell water potential and turgor in osmotic regulation. Potassium content in plants is high compared to trace elements, however, it is difficult to measure a relatively small change of potassium content in the large total. Here, we describe a procedure for ...
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Stem Cell

Serial Transfer of Human Hematopoietic and Hepatic Stem/progenitor Cells Authors:  Qingfeng Chen and Jianzhu Chen, date: 12/05/2013, view: 8512, Q&A: 0
A range of assays have been developed to determine the stemness or stem cell activity of human stem cells. The key assays of stem cells are functional: they must show self-renewal and the ability to generate the appropriate tissue. The best assays available to study this property in putative human stem cells involve xeno-transplantation into ...
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