Mammalia

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    Protocols in Current Issue
    Conjugation of Fab’ Fragments with Fluorescent Dyes for Single-molecule Tracking on Live Cells
    Authors:  I-Ting Teng, Xiangning Bu and Inhee Chung, date: 09/20/2019, view: 214, Q&A: 0
    [Abstract] Our understanding of the regulation and functions of cell-surface proteins has progressed rapidly with the advent of advanced optical imaging techniques. In particular, single-molecule tracking (SMT) using bright fluorophores conjugated to antibodies and wide-field microscopy methods such as total internal reflection fluorescence microscopy have ...
    A Robust, One-step FRET Assay for Human Heparanase
    Authors:  Jyothi C. Sistla and Umesh R. Desai, date: 09/05/2019, view: 277, Q&A: 0
    [Abstract] Heparanase, an endo-β-D-glucuronidase, cleaves cell surface and extracellular matrix heparan sulfate (HS) chains at distinct sites and plays important biological roles including modulation of cell growth and metastasis. Although a number of different types of heparanase assays have been reported to date, most are labor intensive, complex ...
    Cell-based Assay for Recruitment of DDR1 to Collagen-coated Beads
    Authors:  Victoria Juskaite and Birgit Leitinger, date: 08/20/2019, view: 344, Q&A: 0
    [Abstract] The discoidin domain receptors, DDR1 and DDR2, are key signaling receptors for the extracellular matrix protein collagen. The interactions of cells with collagen are difficult to study because of the difficulty to obtain native collagen fibers for in vitro studies. Thus, in vitro studies often use acid-soluble collagens in the ...
    Genotyping of the OATP1B1 c. 521 T>C Polymorphism from the Formalin-Fixed Paraffin-Embedded (FFPE) Tissue Specimens: An Optimized Protocol
    Authors:  Alexandra R Crowe, Jonathan Miller and Wei Yue, date: 08/20/2019, view: 236, Q&A: 0
    [Abstract] Organic anion transporting polypeptide (OATP) 1B1 is a liver-specific transport protein that plays an important role in hepatic drug disposition. It transports many drugs from the blood into the liver, including lipid-lowering statins. The c. 521 T>C polymorphism of OATP1B1 has reduced transport activity and is associated with statin-induced ...
    Electron Microscopy Sample Preparation Protocol Enabling Nano-to-mesoscopic Mapping of Cellular Connectomes and Their Habitats in Human Tissues and Organs
    [Abstract] Multibeam scanning electron microscopy (multiSEM) provides a technical platform for seamless nano-to-mesoscale mapping of cells in human tissues and organs, which is a major new initiative of the U.S. National Institutes of Health. Such cross-length-scale imaging is expected to provide unprecedented understanding of relationships between cellular ...
    Detection of Heteroplasmic Variants in the Mitochondrial Genome through Massive Parallel Sequencing
    Authors:  Joke Mertens, Filippo Zambelli, Dorien Daneels, Ben Caljon, Karen Sermon and Claudia Spits, date: 07/05/2019, view: 695, Q&A: 0
    [Abstract] Detecting heteroplasmies in the mitochondrial DNA (mtDNA) has been a challenge for many years. In the past, Sanger sequencing was the main option to perform this analysis, however, this method could not detect low frequency heteroplasmies. Massive Parallel Sequencing (MPS) provides the opportunity to study the mtDNA in depth, but a controlled ...
    A Detailed Protocol for Large-scale Recombinant Expression and Validation of Human FGFR2 with Its Transmembrane and Extracellular Domains in Escherichia coli
    Authors:  Adam Bajinting and Ho Leung Ng, date: 06/05/2019, view: 770, Q&A: 0
    [Abstract] Receptor tyrosine kinases (RTKs) are an important class of transmembrane receptors that mediate some of the most crucial biochemical pathways essential to the growth, differentiation, and survival of a cell and thus, are highly involved in cancers. Due to the complexity of RTKs having biochemically different domains including a transmembrane ...
    Quantification of Queuosine Modification Levels in tRNA from Human Cells Using APB Gel and Northern Blot
    Authors:  Zaneta Matuszek and Tao Pan, date: 03/20/2019, view: 1083, Q&A: 0
    [Abstract] Queuosine (Q) is a hypermodified base in the wobble anticodon position of tRNAs coding for the amino acids Tyr, His, Asn, and Asp. tRNA Q-modification is introduced by a queuine tRNA-ribosyltransferase (TGT) that replaces the guanine base at G34 at these tRNAs with the modified base. tRNA Q-modification is widely distributed among prokaryotic and ...
    A New Efficient Method for Measuring Oxygen Consumption Rate Directly ex vivo in Human Epidermal Biopsies
    Authors:  Daniel Schniertshauer, Daniel Gebhard and Jörg Bergemann, date: 03/05/2019, view: 1187, Q&A: 0
    [Abstract] Skin cells are constantly exposed to environmental influences such as air pollution, chemicals, pathogens and UV radiation. UV radiation can damage different biological structures, but most importantly cellular DNA. Mitochondria contain their own genome and accumulate UV-induced DNA mutations to a large extent. This can result, e.g., in ...
    Protocol for in situ Proximity Ligation Assay (PLA) and Microscopy Analysis of Epidermal Growth Factor Receptor (EGFR) Homodimerization
    Authors:  Keiichi Ota and Taishi Harada, date: 11/05/2018, view: 2180, Q&A: 0
    [Abstract] Oncogenic drivers play central roles in tumorigenesis as well as in tumor cell survival and proliferation. Mutations of the epidermal growth factor receptor gene (EGFR) that result in constitutive activation of the receptor tyrosine kinase have been identified as oncogenic drivers in a subset of non-small cell lung cancer (NSCLC). ...



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