Biochemistry

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    Protocols in Current Issue
    Isolation of Lipid Rafts from Cultured Mammalian Cells and Their Lipidomics Analysis
    Authors:  Nigora Mukhamedova, Kevin Huynh, Hann Low, Peter J. Meikle and Dmitri Sviridov, date: 07/05/2020, view: 2084, Q&A: 0
    [Abstract] Lipid rafts are distinct liquid-ordered domains of plasma membranes of most eukaryotic cells providing platform for signaling pathways. Lipid composition of rafts is critical for their structural integrity and for regulation of signaling pathways originating from rafts. Here we provide a protocol to isolate lipid rafts from cultured human and ...
    Eicosanoid Isolation from Mouse Intestinal Tissue for ELISA
    Author:  Isabella Rauch, date: 11/05/2018, view: 2600, Q&A: 0
    [Abstract] Activation of inflammasomes in peritoneal macrophages and intestinal epithelial cells (IEC) leads to the release of eicosanoids. To assess the amount of eicosanoids released by IEC, lipids need to be isolated from whole tissue previous to analysis by lipid mass spectrometry or ELISA. This protocol describes how to isolate lipids from intestinal ...
    Crude Preparation of Lipopolysaccharide from Helicobacter pylori for Silver Staining and Western Blot
    Authors:  Hong Li and Mohammed Benghezal, date: 10/20/2017, view: 7281, Q&A: 0
    [Abstract] This protocol provides an easy and rapid method to prepare lipopolysaccharide from the gastric pathogen Helicobacter pylori for visualization on acrylamide gels by silver staining and for detecting the presence of Lewis antigens by Western blot. The silver staining is a four-step procedure, involving a 20 min-oxidation step, a 10 ...
    Sterol Analysis in Kluyveromyces lactis
    Authors:  Yvetta Gbelska, Nora Toth Hervay, Marcela Morvova and Alexandra Konecna, date: 09/05/2017, view: 5153, Q&A: 0
    [Abstract] Sterols are essential lipids of most eukaryotic cells with multiple functions (structural, regulatory and developmental). Sterol profile of yeast cells is often determined during the studies of ergosterol synthesis mutants used to uncover a number of functions for various sterols in yeast cells. Molecular studies of ergosterol biosynthesis have ...
    Preparation of Everted Membrane Vesicles from Escherichia coli Cells
    Author:  Marina Verkhovskaya, date: 05/05/2017, view: 6641, Q&A: 0
    [Abstract] The protocol for obtaining electrically sealed membrane vesicles from E. coli cells is presented. Proton pumps such as Complex I, quinol oxidase, and ATPase are active in the obtained vesicles. Quality of the preparation was tested by monitoring the electric potential generated by these pumps.
    Preparation of Purified Gram-positive Bacterial Cell Wall and Detection in Placenta and Fetal Tissues
    Authors:  Beth Mann, Lip Nam Loh, Geli Gao and Elaine Tuomanen, date: 12/05/2016, view: 7865, Q&A: 0
    [Abstract] Cell wall is a complex biopolymer on the surface of all Gram-positive bacteria. During infection, cell wall is recognized by the innate immune receptor Toll-like receptor 2 causing intense inflammation and tissue damage. In animal models, cell wall traffics from the blood stream to many organs in the body, including brain, heart, placenta and ...
    Determining Efficiency and Selectivity of Lipid Extraction by Perturbing Agents from Model Membranes
    Authors:  Michel Lafleur and Alexandre Therrien, date: 11/20/2016, view: 5999, Q&A: 0
    [Abstract] Several membrane-perturbing agents extract lipids from membranes and, in some cases, this lipid efflux is lipid specific. In order to gain a better description of this phenomenon and to detail the intermolecular interactions that are involved, a method has been developed to characterize the extent and the specificity of membrane-lipid extraction ...
    Lipid Extraction from HeLa Cells, Quantification of Lipids, Formation of Large Unilamellar Vesicles (LUVs) by Extrusion and in vitro Protein-lipid Binding Assays, Analysis of the Incubation Product by Transmission Electron Microscopy (TEM) and by Flotation across a Discontinuous Sucrose Gradient
    [Abstract] Dissecting the interactions established between proteins and membranes in a given type of cells is not an easy task. Using a cell-free system of large unilamellar vesicles (LUVs) to analyze these interactions may help decipher these interactions and identify potential membrane deformations induced by the proteins incubated with these LUVs. This ...
    Extraction and Profiling of Plant Polar Glycerol Lipids
    Authors:  Yu Liu and Xuemin Wang, date: 06/20/2016, view: 9290, Q&A: 0
    [Abstract] This protocol describes a method to extract total polar glycerol lipids from plant materials, followed by mass spectrometry profiling. Different glycerol lipid classes can be distinguished by their head-groups, which can be profiled automatically and quantitatively by a triple quadrupole mass spectrometry in multiple reaction monitoring (MRM) mode ...
    Preparation of Chloroplast Lipid Membrane and Lipid-protein Interaction Assay
    Authors:  Kumiko Okazaki, Shin-ya Miyagishima and Hajime Wada, date: 02/05/2016, view: 7498, Q&A: 0
    [Abstract] Lipid-Protein interaction assay is a method to search lipids, which are bound with proteins in vitro. Since membranes that are spotted with chloroplast lipids such as monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), and sulfoquinovosyldiacylglycerol (SQDG) are not commercially available, we extracted these lipids ...



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