Purification and activity determination of ADAMTS-4 and ADAMTS-5 and their domain deleted mutants by Fowkes, MM, Lim, NH

“…In this chapter we describe the establishment and cultivation of cell lines expressing ADAMTS-4,-5 and their domain deletion mutants; the collection of medium containing expressed ADAMTS-4,-5; the subsequent purification of this medium through anti-FLAG affinity chromatography; and the characterization of ADAMTS-4,-5 activity using synthetic Förster resonance energy transfer (FRET) peptide substrates.…”

Foot-and-mouth disease virus (FMDV) with a stable FLAG epitope in the VP1 G-H loop as a new tool for studying FMDV pathogenesis. by Lawrence, P, Pacheco, J, Uddowla, S, Hollister, J, Kotecha, A, Fry, E, Rieder, E

“…FLAG epitope stability was confirmed both in vitro and in vivo. Interestingly, no anti-FLAG antibodies were detectable in cattle up to 21 days post-inoculation. …”

Positive Selection of Specific Antibodies Produced against Fusion Proteins by Lukas Kramberger-Kaplan, Tina Austerlitz, Holger Bohlmann

“…These proteins were then coupled to NHS-activated sepharose and used for the positive selection of Anti-GFP and Anti-FLAG antibodies. The obtained sera were tested for their specificity against different protein samples and fusion proteins in Western blots. …”
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Generation of Flag/DYKDDDDK Epitope Tag Knock-In Mice Using <i>i</i>-GONAD Enables Detection of Endogenous CaMKIIα and β Proteins by Kazushi Aoto, Shuji Takabayashi, Hiroki Mutoh, Hirotomo Saitsu

“…Experiments with commercially available anti-Flag antibodies could readily detect endogenous CaMKIIα and β proteins by Western blotting, immunoprecipitation, and immunohistochemistry. …”
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Lack of KIR3DS1 binding to MHC class I Bw4 tetramers in complex with CD8+ T cell epitopes. by Gillespie, G, Bashirova, A, Dong, T, McVicar, D, Rowland-Jones, S, Carrington, M

“…Using flow cytometry analysis, the expression of KIR3DS1 on 293-T cells was confirmed by anti-FLAG BioM2 staining, prior to incubation with PE-conjugated MHC class I tetramers. …”

Identification of the ataxin-1 interaction network and its impact on spinocerebellar ataxia type 1 by Jiu-Ming Chen, Shi-Kai Chen, Pei-Pei Jin, Shun-Chang Sun

“…Co-immunoprecipitation was done in HEK-293T cells expressing exogenous wild-type and mutant ataxin-1 using anti-Flag antibody following by tandem affinity purification in order to study protein–protein interactions. …”
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SMR peptide antagonizes Staphylococcus aureus biofilm formation by Ming-Bo Huang, Dara Brena, Jennifer Y. Wu, Martin Shelton, Vincent C. Bond

“…Interaction between the antagonist and DnaK was determined by immune precipitation with anti-Flag M2 Affinity and Western blot analysis. Increasing SMR peptide concentrations exhibited increasing blockade of S. aureus biofilm formation with significant inhibition found at 18 µM, 36 µM, and 72 µM. …”
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Screening of AAK1 interaction proteins and its role in regulating global translation level in cells by JIANG Guixian, HU Ronggui, WU Hao

“…The regulation of AAK1 in the global protein synthesis was explored by puromycin incorporation assay.Results·Mass spectrometry results showed that AAK1 was associated with a series of proteins, including fragile X mental retardation syndrome-related protein 1 (FXR1), FXR2 and fragile X mental retardation protein 1 (FMRP). Enriching with anti-FLAG agarose gels after exogenous transfecting of AAK1-3xFLAG and FMRP-MYC plasmids, the expression of FMRP-MYC was detected. …”
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Generation of two CRISPR/Cas edited human induced pluripotent stem cell lines (DHMi005-A-3 and DHMi005-A-4) carrying a FLAG-tag after exon 9 of the TBX5 gene by H. Lahm, S. Stieglbauer, I. Neb, S.A. Doppler, S. Schneider, E. Dzilic, R. Lange, M. Krane, M. Dreßen

“…The expressed TBX5-FLAG fusion protein can effectively be precipitated by anti-FLAG antibodies. Therefore, these gene-edited iPSC lines represent powerful cellular in vitro tools to unravel TBX5:DNA interactions in detail.…”
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Living-Cell Diffracted X-ray Tracking Analysis Confirmed Internal Salt Bridge Is Critical for Ligand-Induced Twisting Motion of Serotonin Receptors by Kazuhiro Mio, Shoko Fujimura, Masaki Ishihara, Masahiro Kuramochi, Hiroshi Sekiguchi, Tai Kubo, Yuji C. Sasaki

“…The 5-HT_2ARs were transiently expressed in HEK 293 cells, and the gold nanocrystals were attached to the N-terminal introduced FLAG-tag via anti-FLAG antibodies. The motions were recorded with a frame rate of 100 μs per frame. …”
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Translational profiling identifies sex-specific metabolic and epigenetic reprogramming of cortical microglia/macrophages in APPPS1-21 mice with an antibiotic-perturbed-microbiome by Shabana M. Shaik, Yajun Cao, Joseph V. Gogola, Hemraj B. Dodiya, Xulun Zhang, Hejer Boutej, Weinong Han, Jasna Kriz, Sangram S. Sisodia

“…Methods Six groups of mice that included WT-CD11br, antibiotic (ABX) or vehicle-treated APPPS1-21-CD11br males and females were sacrificed at 7-weeks of age (n = 15/group) and used for immunoprecipitation of microglial/macrophage polysomes from cortical homogenates using anti-FLAG antibody. Liquid chromatography coupled to tandem mass spectrometry and label-free quantification was used to identify newly synthesized peptides isolated from polysomes. …”
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Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization by Bright D. Danquah, Kwabena F. M. Opuni, Claudia Roewer, Cornelia Koy, Michael O. Glocker

“…For the epitope peptide dissociation reaction of the FLAG peptide from the antiFLAG antibody complex an apparent gas phase dissociation constant <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.04 × 10⁻¹² was calculated. …”
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Insertion of a FLAG-tag sequence at the end of exon 9 of the TBX5 gene in three induced pluripotent stem cell lines (DHMi004-A-4, DHMi004-A-5, DHMi004-A-6) by CRISPR/Cas9 technolog... by Harald Lahm, Niraj K. Singh, Irina Gottmann, Stefanie A. Doppler, Elda Dzilic, Heike Preisler, Stephanie Schneider, Rüdiger Lange, Markus Krane, Martina Dreßen

“…The translated TBX5-FLAG fusion protein of the three iPSC lines can effectively be precipitated by anti-FLAG antibodies and, thus, allow the detection of specific TBX5-binding sites and their associated genes.…”
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