Mammalia

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    Protocols in Current Issue
    Extraction and Analysis of Pan-metabolome Polar Metabolites by Ultra Performance Liquid Chromatography–Tandem Mass Spectrometry (UPLC-MS/MS)
    Authors:  Dania M. Malik, Seth Rhoades and Aalim Weljie, date: 02/05/2018, view: 3375, Q&A: 0
    [Abstract] Modern triple quadrupole mass spectrometers provide the ability to detect and quantify a large number of metabolites using tandem mass spectrometry (MS/MS). Liquid chromatography (LC) is advantageous, as it does not require derivatization procedures and a large diversity in physiochemical characteristics of analytes can be accommodated through a ...
    The RiboPuromycylation Method (RPM): an Immunofluorescence Technique to Map Translation Sites at the Sub-cellular Level
    Authors:  Amandine Bastide, Jonathan W. Yewdell and Alexandre David, date: 01/05/2018, view: 5080, Q&A: 0
    [Abstract] While isotopic labeling of amino acids remains the reference method in the field for quantifying translation rate, it does not provide any information on spatial localization of translation sites. The rationale behind developing the ribopuromycylation method (RPM) was primarily to map translation sites at the sub-cellular level while avoiding ...
    GC/MS-based Analysis of Volatile Metabolic Profile Along in vitro Differentiation of Human Induced Pluripotent Stem Cells
    [Abstract] Human induced pluripotent stem cells (hiPSCs) are a promising tool in cell-based therapies for degenerative diseases. A safe application of hiPSCs in vivo, requires the detection of the presence of residual undifferentiated pluripotent cells that can potentially cause the insurgence of teratomas. Several studies point out that metabolic ...
    DQ-Red BSA Trafficking Assay in Cultured Cells to Assess Cargo Delivery to Lysosomes
    Authors:  Rituraj Marwaha and Mahak Sharma, date: 10/05/2017, view: 7323, Q&A: 0
    [Abstract] Lysosomes are the terminal end of the endocytic pathway having acidic environment required for active hydrolases that degrade the cargo delivered to these compartments. This process of cargo delivery and degradation by endo-lysosomes is a tightly regulated process and important for maintaining cellular homeostasis. Cargos like EGF (Epidermal ...
    Quantitative Analysis of Exosome Secretion Rates of Single Cells
    Authors:  Yu-Jui Chiu, Wei Cai, Tiffany Lee, Julia Kraimer and Yu-Hwa Lo, date: 02/20/2017, view: 7279, Q&A: 0
    [Abstract] To study the inhomogeneity within a cell population including exosomes properties such as exosome secretion rate of cells and surface markers carried by exosomes, we need to quantify and characterize those exosomes secreted by each individual cell. Here we develop a method to collect and analyze exosomes secreted by an array of single cells using ...
    Quantification of 2-Hydroxyglutarate Enantiomers by Liquid Chromatography-mass Spectrometry
    Authors:  William M. Oldham and Joseph Loscalzo, date: 08/20/2016, view: 6512, Q&A: 0
    [Abstract] Two enantiomers of 2-hydroxyglutarate (2HG), L (L2HG) and D (D2HG), are metabolites of unknown function in mammalian cells that were initially associated with separate and rare inborn errors of metabolism resulting in increased urinary excretion of 2HG linked to neurological deficits in children (Chalmers et al., 1980; Duran et al ...
    [14C]-Tryptophan Metabolic Tracing in Liver Cancer Cells
    Authors:  Krishna S. Tummala and Nabil Djouder, date: 09/05/2015, view: 5571, Q&A: 1
    [Abstract] Nicotinamide adenine dinucleotide (NAD+) is a coenzyme for many NAD+-consuming proteins with diverse biological functions. Oscillations in NAD+ levels may influence several cellular signaling pathways. NAD+ synthesis via Preiss-Handler route (salvage reactions) has been extensively reported. However, the ...
    Measurement of Acetylcholine from Cell Lines
    Authors:  Jamie K. Lau, Kathleen C. Brown and Piyali Dasgupta, date: 12/20/2013, view: 7246, Q&A: 0
    [Abstract] Cigarette smoking is the leading risk factor for the development of lung cancer. It is estimated that smoking is associated with 80-90% of lung cancer cases throughout the world (see References 1 and 2). The addictive component of cigarette smoke is nicotine. Our published data shows that nicotine promotes the production of acetylcholine (ACh) in ...
    Flow Cytometric Analysis of Calcium Influx Assay in T cells
    Author:  Sun-Hwa Lee, date: 09/20/2013, view: 14328, Q&A: 0
    [Abstract] Calcium influx is one of the key signaling events upon stimulation of T cell receptors (TCR) and plays an important role for T cell activation, proliferation, and differentiation. Phorbol myristate acetate (PMA) and calcium ionophore ionomycin are commonly utilized as stimulants in a variety of immunologic assays including T cell activation. PMA ...
    Sodium Current Measurements in HEK293 Cells
    [Abstract] This protocol is used to measure sodium currents from heterologously transfected cells lines, such as HEK293 cells. Standard whole cell patch clamp technique is used to assess ion channel function. This protocol has been used to study cardiac type Nav1.5 sodium channel, and in particular to compare wild-type and mutant channels related ...



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