This protocol describes a cost effective method for generating highly concentrated yeast whole cell extracts (WCE) at concentrations of 50 μg protein/μl or 0.3 A260nm/µl. Highly concentrated extracts are used for many applications such as protein purification, purification of protein complexes (in particular when interactions are weak), protein-protein interaction studies, and velocity sediment...

Authors: Jyothsna Visweswaraiah, Martina Dautel and Evelyn Sattlegger
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Antimicrobial peptides represent the first-line host defence against microbial pathogens and an essential component of innate immunity. They have received growing interest because of their potential use as therapeutic antibiotics. Due to the fact that most antimicrobial peptides are toxic to prokaryotic host cells, they are currently often produced by chemical synthesis. However, this is too costl...

Authors: Zhihong Wu et al
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Postoperative ileus (POI) affects patients after abdominal surgery. Standardized rodent models of intestinal manipulation represent the surgical trauma. POI is induced by local inflammation in the intestinal muscularis. This inflammation is due to an activation of resident muscularis macrophages leading to paralysis of intestinal myocytes.This disseminates along the entire gastrointestinal tract, ...

Authors: Arne Koscielny et al
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Organisms store excess energy as fatty acids and triglycerides. Under fasting conditions, fatty acids are oxidized in the mitochondria in a multi-step pathway called beta-oxidation1. Beta-oxidation (or fatty acid oxidation) begins by importing long-chain fatty acids into the mitochondria, followed by a four-step reaction. In the first step, acyl-CoA dehydrogenase dehydrogenates the long-chain acyl...

Authors: Matthew D Hirschey and Eric Verdin
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Given that DNA on which genomic information is written exists as chromosomes in a cell, handling chromosomes in vitro as experimental materials can provide varieties of information throughout life sciences. Metaphase chromosomes are highly delicate under in vitro conditions, moreover, it has been difficult to prepare massive chromosomes as experimental materials. These inconvenient points have pre...

Authors: Kayoko Hayashihara et al
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The Guava Easy-Cyte flow cytometer accepts 96 well plates containing stained cells in liquid media and reports the cells/ml for each well in Excel format (csv file). Marine microbes stained with SYBR-Grn I in natural seawater can be counted within approximately 5% error at densities of 1.0E4 to 5.5E5 cells per ml. With exceptional care, cell cultures can be reliably detected but not accurately cou...

Authors: H Jams Tripp
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Processing urine samples in this way allows urine to be used for several proteomic¨applications, including 1D and 2D-PAGE and SELDI/MALDI profiling. The effect of addition of protease inhibitor tablets is currently being evaluated in matched samples (+/- PI) from patients with either renal cancer, post-renal transplant or healthy controls, stored for various periods of time. If necessary the pro...

Authors: Roz Banks
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We (at Inserm U858 Toulouse, France) have prepared rat- and mouse kidneys for the LCM training school in Rotterdam, NL (May 25-27, 2009). The feedback was that these samples obtained by a simple protocol were suitable for LCM-proteomics on Hematoxylin stained sections. The following protocol was used for both mouse (17 weeks old male C57Bl/6J strain) and rat (3 months old female Sprague Dawley) ki...

Authors: Joost Schanstra
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This is a general protocol for urine collection known to provide overall good proteomics data quality for a wide spectrum of technologies and diseases under investigation, based on our accumulated knowledge so far. However it should be noted that for special cases/diseases (eg associated with high proteinuria, hematuria etc) the application of specialized protocols may be recommended. It is likel...

Authors: EU COST action
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In order to facilitate the generation of meaningful data and to clarify the requirements for the analysis, we have prepared this sheet, indicating the most important steps.

Authors: Joint Protocol
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