• Volume 9, 2019
  • Volume 8, 2018
  • Volume 7, 2017
  • Volume 6, 2016
  • Volume 5, 2015
  • Volume 4, 2014
  • Volume 3, 2013
  • Volume 2, 2012
  • Volume 1, 2011

Cancer Biology

Lysosomal Stability Assay Authors:  Nikolaj H. T. Petersen, Thomas Kirkegaard and Marja Jäättelä, date: 06/20/2014, view: 11859, Q&A: 0
This assay makes use of the dye Acridine Orange (AO) to determine the stability of lysosomes in living cells upon exposure to a confocal microscope laser.

AO is a lipophilic amine that readily diffuses into cells (Figure 1). Inside the cell it enters the acidic lysosomal compartment where it is protonated and sequestered, shifting its ...
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Cell Biology

EML Erythroid and Neutrophil Differentiation Protocols Authors:  Cristina Pina, Cristina Fugazza and Tariq Enver, date: 06/20/2014, view: 7812, Q&A: 0
Erythroid-Myeloid-Lymphoid cells (EML) are a multipotent haematopoietic cell line of mouse bone marrow origin capable of long-term maintenance in vitro in the presence of SCF (stem cell factor) (Tsai et al., 1994). The self-renewal capacity of the EML cell line is conferred by the presence of a dominant-negative retinoic acid ...
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Immunology

Bone Marrow Derived Eosinophil Cultures Authors:  Thomas X. Lu and Marc E. Rothenberg, date: 06/20/2014, view: 9231, Q&A: 1
Eosinophils are multifunctional effector cells implicated in the pathogenesis of a variety of diseases including asthma, eosinophil gastrointestinal disorders and helminth infection. Mouse bone marrow derived progenitor cells can be differentiated into eosinophils following IL-5 exposure. These bone marrow derived eosinophils are fully ...
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Microbiology

A SYBR Green-based Real Time RT-PCR Assay for Detection of the Emerging H7N9 Virus Authors:  Zheng Zhu and Lunbiao Cui, date: 06/20/2014, view: 6076, Q&A: 0
Most recently a novel avian-origin influenza A (H7N9) virus emerged in China and has been associated with lots of human infection and fatal cases. Molecular diagnostic methods are thus urgently needed in public health laboratories. We developed a SYBR green-based one-step real time reverse transcription-PCR (RT-PCR) to detect the novel H7N9 virus.
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Determination of Oxygen Respiration Rates in Wetted Developmentally Arrested Spores of Streptomyces Species Authors:  Marco Fischer, Dörte Falke and R. Gary Sawers, date: 06/20/2014, view: 5977, Q&A: 0
Streptomyces species produce spores, which, while not as robust as endospores of Bacillus or Clostridium species, are capable of surviving for months or even years (Hopwood, 2006). During this time these spores remain viable, surviving by slowly degrading internal stores of carbon compounds, such as the carbohydrate ...
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Rapid Nitrate Reduction Assay with Intact Microbial Cells or Spores Authors:  Marco Fischer, Dörte Falke and R. Gary Sawers, date: 06/20/2014, view: 5924, Q&A: 0
Many microorganisms have the capacity to use nitrate as a respiratory electron acceptor. Reduction of nitrate is catalyzed by a multi-subunit nitrate reductase that is often located associated with the cytoplasmic membrane and has its active site oriented toward the cytoplasm. This means that nitrate must be transported into the cell and often ...
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Analysis of Mycobacterial Protein Secretion Authors:  Alka Mehra and Jennifer A. Philips, date: 06/20/2014, view: 9356, Q&A: 0
Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis. Analysis of proteins secreted by Mtb has been of interest to the field of tuberculosis research since certain secreted proteins interact with the host to promote virulence, while others may be important antigens or serve as biomarkers of infection. Here, we describe ...
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Plant Science

Protoplast Preparation and Determination of Cell Death Author:  Antoine Danon, date: 06/20/2014, view: 11651, Q&A: 3
The protoplasts assay constitutes a powerful tool that allows an easy uptake of active agents and a precise quantification of cell death induction in different populations. Our study showed that the basal level of cell death in our controls is low and stable throughout the length of our experiments (Danon et al., 2005; Pineau et al. ...
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MTV1 Pull-down Assay in Arabidopsis Author:  Michael Sauer, date: 06/20/2014, view: 7825, Q&A: 0
This protocol is an example of how to analyse suspected interactions between proteins using a pull-down assay (Sauer et al., 2013). A bait protein of interest (in this case, MTV1 of Arabidopsis thaliana) is fused to a GST tag and expressed in bacteria. The protein is isolated and allowed to bind to a matrix of ...
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Deflagellation and Regeneration in Chlamydomonas Author:  William Dentler, date: 06/20/2014, view: 10226, Q&A: 0
Eukaryotic cilia/flagella are one of the only cellular structures that can be removed without injuring cells, can be highly purified for biochemical analysis, and, in many cells, can be completely reassembled within 90 minutes. Following amputation, the expression of many flagellar genes is up-regulated, and many are packaged and associated with ...
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Ciliary and Flagellar Membrane Vesicle (Ectosome) Purification Author:  William Dentler, date: 06/20/2014, view: 6971, Q&A: 0
Eukaryotic cilia/flagella are ideal organelles for the analysis of membrane trafficking, membrane assembly, and the functions of a variety of signal transduction molecules. Cilia are peninsular organelles and the membrane lipids, membrane proteins, and microtubular-associated components are selectively transported into cilia through the region ...
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In organello Protein Synthesis Authors:  Malgorzata Kwasniak-Owczarek and Hanna Janska, date: 06/20/2014, view: 8170, Q&A: 1
In organello protein synthesis method allows the analysis of mitochondrial translation products. The principle of this method relies on incubation of isolated intact mitochondria with radiolabeled amino acids such as 35S methionine. After protein synthesis, the radiolabeled translation products are subsequently separated by SDS ...
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Determination of Luciferase Activity in Arabidopsis seedling Authors:  Mohan TC, Gabriel Castrillo and Antonio Leyva, date: 06/20/2014, view: 8339, Q&A: 0
There are different direct and indirect methods available to study gene expression in plant systems. In this protocol we describe a modified expression assay using transgenic plants expressing the luciferase gene under the control of phosphate transporter PHT1;1 promoter. This assay was originally optimized for analyzing the repression of PHT1;1 ...
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Systems Biology

ChIP-Seq in Candida albicans Authors:  Sadri Znaidi, Caroline Proux, Sandra Weber, Simon Drouin, François Robert, Martine Raymond, Jean-Yves Coppée and Christophe d’Enfert, date: 06/20/2014, view: 12005, Q&A: 0
Systems biology approaches can be used to study the regulatory interactions occurring between many components of the biological system at the whole-genome level and decipher the circuitries implicated in the regulation of cellular processes, including those imparting virulence to opportunistic fungi. Candida albicans (C. albicans ...
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