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Immunology

Binding Affinity Measurement of Antibodies from Crude Hybridoma Samples by SPR Authors:  Dorin Mlaki Ndao, David T. Hickman, María Pilar López-Deber, Aurélien Davranche, Andrea Pfeifer and Andreas Muhs, date: 11/05/2014, view: 7364, Q&A: 0
Surface Plasmon Resonance (SPR) is widely used to generate kinetic and affinity information on specific interactions between biomolecules. This technique is label-free and monitors the binding event in real-time. It is generally used for characterization of monoclonal antibody - antigen interactions. This protocol describes specifically the use of ...
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Isolation and in vitro Activation of Mouse Peyer’s Patch Cells from Small Intestine Tissue Authors:  Claudia Pastori and Lucia Lopalco, date: 11/05/2014, view: 7576, Q&A: 0
The lumen of gastrointestinal tract is exposed to several potentially pathogenic microorganisms, thus it is extremely relevant to understand how immunosurveilance can be established. Peyer’s Patches (PPs) are oval or round lymphoid nodules that protrude from the outer wall of the ileum portion of small intestine. PPs contain a high percentage of B ...
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Microbiology

Measurement of Proton-driven Antiport in Escherichia coli Authors:  Scarlett R. Holdsworth and Christopher J. Law, date: 11/05/2014, view: 4060, Q&A: 0
Secondary active transport of substrates across the inner membrane is vital to the bacterial cell. Of the secondary active transporter families, the ubiquitous major facilitator superfamily (MFS) is the largest and most functionally diverse (Reddy et al., 2012). Recently, it was reported that the MFS multidrug efflux protein MdtM from ...
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Measurement of the Electrogenicity of Bile Salt/H+ Antiport in Escherichia coli Authors:  Scarlett R. Holdsworth and Christopher J. Law, date: 11/05/2014, view: 3564, Q&A: 0
The transmembrane proton gradient (ΔpH) is the primary source of energy exploited by secondary active substrate/H+ antiporters to drive the electroneutral transport of substrates across the Escherichia coli (E. coli) inner membrane. Such electroneutral transport results in no net movement of charges across the ...
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Purification and Detection of a PDGA Depolymerase from Pusillimonas noertemannii Authors:  David Negus and Peter W. Taylor, date: 11/05/2014, view: 4072, Q&A: 0
The purification of a target protein from a complex mixture of proteins is a challenging undertaking. If the target protein has been previously characterised, then information such as subcellular location, function, molecular weight and pI can be used for the design of a purification strategy. However, if the target protein is uncharacterised or ...
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Accelerated Storage Stability Testing of a Potential Anti-Anthrax Therapeutic, EnvD Authors:  David Negus and Peter W. Taylor, date: 11/05/2014, view: 3534, Q&A: 0
The purpose of stability testing is to determine how the properties of a particular therapeutic vary with time under the influence of specific environmental factors. Information regarding the long-term stability of therapeutics can be extrapolated by performing an accelerated storage stability study. Here, we describe an accelerated storage ...
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Stable Transformation of Cyanobacterium Synechocystis sp. Author:  Reinhard K Proels, date: 11/05/2014, view: 6200, Q&A: 0
Cyanobacteria are prokaryotes, which perform oxygenic photosynthesis. Among them, the unicellular cyanobacterium Synechocystis sp. PCC 6803 (hereafter Synechocystis) is a well characterized model system for studies on oxygenic photosynthesis, light signal transduction etc. Moreover, Synechocystis is applied in ...
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Fractionation by Ultracentrifugation of Gram Negative Cytoplasmic and Membrane Proteins Authors:  Sara M Sandrini, Richard Haigh and Primrose P. E. Freestone, date: 11/05/2014, view: 9314, Q&A: 1
Protein fractionation is a useful separation process which divides membrane proteins (including those located in the outer and inner membrane) and cytoplasmic proteins into discrete fractions. Fractionation of proteins can simplify analysis of the numbers of proteins present, and therefore make easier to characterize any environmentally or ...
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Neuroscience

An Experimental Model of Neonatal Nociceptive Stimulation in Rats Authors:  Jackeline Moraes Malheiros, Cristiane Amaral, Ana Teresa S Leslie, Ruth Guinsburg and Luciene Covolan, date: 11/05/2014, view: 4894, Q&A: 0
In order to survive, preterm and/or sick neonates need diagnostic and therapeutic measures that may cause discomfort, stress and pain during a critical period of intense growing and modeling of the central nervous system (Anand et al., 2013). Scientific interest in the long lasting effects of the Neonatal Intensive Care (NIC) experience, ...
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Pain Assessment Using the Rat and Mouse Formalin Tests Authors:  Nian Gong, Qian Huang, Yuan Chen, Meng Xu, Shuai Ma and Yong-Xiang Wang, date: 11/05/2014, view: 11995, Q&A: 0
The formalin test was originally developed by Dubuisson and Dennis (1977), and has since been extensively used to assess pain-related responses. Rats and mice are the most frequently used animal models, though other species including cats, rabbits, guinea pigs, Octodon degus, domestic fowls, crocodiles, tortoises, toads and primates have ...
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Plant Science

Preparation of Multiplexed Small RNA Libraries from Plants Authors:  Kerrigan B. Gilbert, Noah Fahlgren, Kristin D. Kasschau, Elisabeth J. Chapman, James C. Carrington and Alberto Carbonell, date: 11/05/2014, view: 7006, Q&A: 0
High-throughput sequencing is a powerful tool for exploring small RNA populations in plants. The ever-increasing output from an Illumina Sequencing System allows for multiplexing multiple samples while still obtaining sufficient data for small RNA discovery and characterization. Here we describe a protocol for generating multiplexed small RNA ...
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Extraction of Total Proteins from Rice Plant Authors:  Da-Gin Lin and Chang-Sheng Wang, date: 11/05/2014, view: 16019, Q&A: 0
This protocol provides an efficient method for preparation of high-quality proteins from rice leaves and grains. The method involves phenol extraction to separate proteins from the non-protein components such as polysaccharides, lipids and phenolic compounds that are commonly enriched in plant tissues. Following isolation, proteins are ...
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Measurement of the Number of Peroxisomes Authors:  Michitaro Shibata, Kazusato Oikawa, Shoji Mano and Mikio Nishimura, date: 11/05/2014, view: 4500, Q&A: 0
This is the detailed protocol for the measurement of the number of peroxisomes described by Shibata et al. (2013). It is difficult to count the number of organelles in a cell because of the thickness of plant leaves. To overcome this challenge, protoplasts were isolated from leaves, and the number of peroxisomes per protoplast was ...
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In vitro Assay of the Glycosyltransferase Activity of a Heterologously Expressed Plant Protein Authors:  Carsten P Ade, Felix Bemm, James M J Dickson, Christian Walter and Philip J Harris, date: 11/05/2014, view: 5524, Q&A: 0
Glycosyltransferases are carbohydrate active enzymes containing catalytic modules involved in catalysing the biosynthesis of glycosidic bonds in oligo- and polysaccharides and glycoconjugates. One of the most comprehensive collections of Carbohydrate Active enZYmes is the CAZy database (http://www.cazy.org ...
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