Molecular Biology

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    Proteome Integral Solubility Alteration (PISA) Assay in Mammalian Cells for Deep, High-Confidence, and High-Throughput Target Deconvolution

    Chemical proteomics focuses on the drug–target–phenotype relationship for target deconvolution and elucidation of the mechanism of action—key and bottleneck in drug development and repurposing. Majorly due to the limits of using

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    In-Cell Western Protocol for Semi-High-Throughput Screening of Single Clones
    Authors:  Arpita S. Pal, Alejandra M. Agredo and Andrea L. Kasinski, date: 08/20/2022, view: 1451, Q&A: 0

    The in-cell western (ICW) is an immunocytochemical technique that has been used to screen for effects of siRNAs, drugs, and small molecule inhibitors. The reduced time and number of cells required to follow this protocol illustrates its

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    A High-throughput Automated ELISA Assay for Detection of IgG Antibodies to the SARS-CoV-2 Spike Protein

    The SARS-CoV-2 pandemic and vaccination campaign has illustrated the need for high throughput serological assays to quantitatively measure antibody levels. Here, we present a protocol for a high-throughput colorimetric ELISA assay to detect IgG

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    Optimised Method for the Production and Titration of Lentiviral Vectors Pseudotyped with the SARS-CoV-2 Spike

    The use of recombinant lentivirus pseudotyped with the coronavirus Spike protein of SARS-CoV-2 would circumvent the requirement of biosafety-level 3 (BSL-3) containment facilities for the handling of SARS-CoV-2 viruses. Herein, we describe a fast

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    Differential Analysis of N-glycopeptide Abundance and N-glycosylation Site Occupancy for Studying Protein N-glycosylation Dysregulation in Human Disease
    Authors:  Qi Zhang, Cheng Ma, Lian Li and Lih-Shen Chin, date: 06/20/2021, view: 2436, Q&A: 0

    Protein N-glycosylation plays a vital role in diverse cellular processes, and dysregulated N-glycosylation is implicated in a variety of human diseases including neurodegenerative disorders and cancer. With recent advances in high-resolution mass

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    In vitro Measurement of Membrane Attack Complex in RPE Cells
    Authors:  Kelly Mulfaul and Sarah L. Doyle, date: 02/20/2021, view: 2926, Q&A: 0

    Initiation of the complement system results in the formation of a multiprotein pore termed the membrane attack complex (MAC, C5b-C9). MAC pores accumulate on a cell surface and can result in cell lysis. The retinal pigment epithelium (RPE) is a

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    Assembly and Imaging Set up of PIE-Scope
    Cryo-Electron Tomography (cryo-ET) is a method that enables resolving the structure of macromolecular complexes directly in the cellular environment. However, sample preparation for in situ Cryo-ET is labour-intensive and can require both ...
    Quantification of the Surface Expression of G Protein-coupled Receptors Using Intact Live-cell Radioligand Binding Assays
    Authors:  Xin Xu and Guangyu Wu, date: 09/20/2020, view: 3260, Q&A: 0
    G protein-coupled receptors (GPCRs) are the most structurally diverse family of signaling proteins and regulate a variety of cell function. For most GPCRs, the cell surface is their functional destination where they are able to respond a wide range ...
    A Workflow for Ultra-rapid Analysis of Histone Post-translational Modifications with Direct-injection Mass Spectrometry
    Authors:  Natarajan V. Bhanu, Simone Sidoli and Benjamin A Garcia, date: 09/20/2020, view: 3912, Q&A: 0
    Chromatin modifications, like histone post translational modifications (PTMs), are critical for tuning gene expression and many other aspects of cell phenotype. Liquid chromatography coupled to mass spectrometry (LC-MS) has become the most suitable ...
    Immunophenotyping and Intracellular Staining of Fixed Whole Blood for Mass Cytometry (CyTOF)
    Authors:  Sangeeta Kowli and Holden Maecker, date: 09/05/2020, view: 4882, Q&A: 1
    In this report, we present the implementation of mass cytometry for intracellular staining using fixed whole blood. In our assay described here, 250 µl of whole blood, is stimulated in vitro with PMA/ionomycin (or left unstimulated), in the ...



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