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High sensitive detection of follistatin by Imperacer®: an initial study on the way to analytical gene-doping tests
For the understanding of interactions in the muscle-growth directing myostatin-pathway and therefore also for the establishment of high sensitive detection strategies for gene-doping, detailed studies of protein concentration levels in biological matrices and appropriate highly sensitive analytic techniques for quantitative monitoring of these biomarkers are required. In this work, the development of a high sensitive assay for follistatin based on the Imperacer® technology is described. The quantification limit in human serum was found at 60 pg/ml in only 6 µl sample volume. With this initial assay we demonstrate the first application in a set of analytical tests for a high sensitive myostatin pathway related profiling....
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Molecular Probes® NanoOrange® Assay Performed on BMG LABTECH FLUOstar OPTIMA Microplate Reader
The field of proteomics has expanded dramatically in recent years with research on a whole host of organisms. Techniques such as two dimensional difference gel electrophoresis (2D DIGE) require the accurate quantitation of protein, as up to three labelled samples can be loaded on a single gel for comparison. Assay methods for determining protein quantitation include absorbance at 280 nm(1), the Bradford assay(2), Lowry assay(3), BCA method(4) and more sensitive assays such as Fluoroprofile® (Sigma-Aldrich) and NanoOrange® (Molecular Probes®)5()....
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Bradford Assay Performed on BMG LABTECH“s FLUOstar Omega with new Evaluation Software
Determining the protein concentration of samples is a necessary and often used method in biochemistry. Different colorimetric protein assays have been developed. The most commonly used methods are the Bradford assay, the Lowry assay and the BCA assay. In this application note we demonstrate how to determine the protein concentration of samples by using the Bradford assay and the new FLUOstar Omega....
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Efficient Transfection of Primary Human Skeletal Myoblasts Using FuGENE® HD Transfection Reagent
Transfection of cells is one of the main techniques used to influence gene expression. Most primary cells and human skeletal myoblasts (SkMC) in particular are very difficult to transfect, whereas for cell lines such as C2C12, many suitable transfection reagents and protocols are available. Few publications report successful transfection of human primary myoblasts using non-viral systems [1,2,3]. These methods include cationic lipids such as phosphonolipids, electroporation, and a combination of liposome and adenoviral associated proteins. But transfection efficiency is low and often is a compromise between toxicity of the reagents and transfection efficiency....
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Enzymatic Protein Digest for Mass-spectrometric Analysis
The main focus in proteomic studies is on the identification of proteins in given biological samples. Proteins isolated and separated from a given sample (e.g., whole cell lysates, blood or tissue, protein complexes) by immunoprecipitation or affinity chromatography or two-dimensional electrophoresis must be proteolytically cleaved in the course of sample preparation for identification and characterization. A reproducible cleavage pattern of digested proteins is a prerequisite for the unambiguous identification of these proteins by mass spectrometry (MS)....
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