Flag f7425

Manufactured by Merck Group

Sourced in United States

The FLAG (F7425) is a laboratory equipment product manufactured by the Merck Group. It serves as a tool for research and scientific applications. The core function of the FLAG (F7425) is to provide a reliable and standardized method for the detection and purification of tagged proteins. The product specifications and technical details are available upon request.

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Lab products found in correlation

10 protocols using flag f7425

Molecular and Cellular Characterization of HCV Infection

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General materials were purchased from Sigma-Aldrich or VWR. DMEM, FBS, Opti-MEM, and Lipofectamine 3000 were purchased from Thermo Fisher Scientific. Protease inhibitor cocktail (P8340; Sigma-Aldrich) was used at 1:100 dilution. Monoclonal antibody C7-50 to HCV core protein and monoclonal antibody JL-8 to GFP were purchased from Thermo Fisher Scientific. RILP (H85), ApoE (A1.4), CD63 (MX-49.129.5), CD81 (5A6), and GAPDH (FL-335) were purchased from Santa Cruz Biotechnology. Rab7 (D95F2), histone H2B (2722), β-Tubulin, Flotillin-1 (D2V7J), Argonaute 2 (C34C6), Hrs (D7T5N), NLRP3 (D4D8T), FMRP (4317S), GM130 (D6B1), Tom20 (D8T4N), and Caspase-4 (4450) were purchased from Cell Signaling Technology. FMR1 (F4055) and FLAG (F7425) were purchased from Sigma-Aldrich. Caspase-1 (EPR19675), IL-1β (EPR21086), TSG101 (EPR7130(B)), gasdermin D (EPR19829), and HA (HA.C5) were purchased from Abcam. Lamp1 (H4A3) was purchased from Developmental Studies Hybridoma Bank.

Wozniak A.L., Adams A., King K.E., Dunn W., Christenson L.K., Hung W.T, & Weinman S.A. (2020). The RNA binding protein FMR1 controls selective exosomal miRNA cargo loading during inflammation. The Journal of Cell Biology, 219(10), e201912074.

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Metabolic Profiling of Cellular Signaling

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GLUT-1 (ab652) and H3K27Ac (ab4729) were purchased from Abcam. HK (A0994) and horseradish peroxidase (HRP)-goat anti-Rabbit were purchased from ABclonal. HRP-conjugate anti-mouse (w402b) was purchased from Promega. H2BK (07-373), H4K (06-866), and H3K (06-599) were purchased from Millipore. Flag (F7425) was purchased from Sigma-Aldrich. All other antibodies including Rictor (2140), PRAS40 (2691), AKT (9272), GSK3b (9315), ACC (3676), ACLY (4332), FASN (3180), PPARg (2443), ACSS2 (3658), S6 (2217), S473-AKT (4058), T308-AKT (4056), S455-ACLY (4331), T246-pPRAS40 (2997), HA-tag (2367), S9-GSK3b (9315), S235/236-S6 (4858), H3 (9715), H3K9Ac (9649), H3K14Ac (7627), and a-Tubulin (2125) were purchased from Cell Signaling Technologies. 4-OHT was obtained from Toronto Research Chemicals. D-[U-¹⁴C]-glucose (NEC042V250UC) was purchased from Perkin Elmer. MK2206 (S1078) was purchased from Selleck Chemicals. T3 (T2877), dexamethasone (D1756), IBMX (I5879), and insulin (I2643) were from Sigma-Aldrich. [U-¹³C] glucose (CLM-1396-1) and [1,2-¹³C] acetate (CLM-440-PK) were purchased from Cambridge Isotope Laboratories, Inc.

Martinez Calejman C., Trefely S., Entwisle S.W., Luciano A., Jung S.M., Hsiao W., Torres A., Hung C.M., Li H., Snyder N.W., Villén J., Wellen K.E, & Guertin D.A. (2020). mTORC2-AKT signaling to ATP-citrate lyase drives brown adipogenesis and de novo lipogenesis. Nature Communications, 11, 575.

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Antibody Reagents for MINERVA Detection

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The primary antibody against MINERVA (5122) was purchased from Cell Signaling Technology (Danvers, MA, USA) or from NOVUSBIO (NBP1-88783) for western blot or immunofluorescence; those against β-actin (A300-491A) and HA (A190-108A) were purchased from Bethyl Laboratories (Montgomery, TX, USA); those against FLAG M2 (F1804) and FLAG (F7425) were purchased from Sigma-Aldrich (St. Louis, MO, USA) for western blot; those against FLAG (D6W5B) were purchased from Cell Signaling Technology (Danvers, MA, USA) for immunofluorescence. The secondary antibodies, horseradish peroxidase (HRP)-linked anti-rabbit (A120-101P) and anti-mouse (A90-116P), were purchased from Bethyl Laboratories (Montgomery, TX, USA). Lipofectamine 2000 (11668019) and RNAiMAX (13778150) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).

Hahn H., Lee D.E., Jang D.M., Kim J., Lee Y., Cheong H., Han B.W, & Kim H.S. (2020). Structural Insight on Functional Regulation of Human MINERVA Protein. International Journal of Molecular Sciences, 21(21), 8186.

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Antibody Immunofluorescence and Western Blot

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Mouse monoclonal antibodies: acetylated Tubulin (T7451, IF: 1:10,000; Sigma-Aldrich), ARL13B (66739-1-Ig, IF: 1:300; Proteintech, 75-287, IF: 1:1,000; NeuroMab), and S-tag (MAC112, IF: 1:100, WB: 1:5,000; EMD Millipore). Rat monoclonal antibody: HA (7c9, WB: 1:1,000; ChromoTek). Rabbit polyclonal antibodies: EGFP (50403-2-AP, IF: 1:200, WB: 1:1,000; Proteintech), Flag (F7425, WB: 1:1,000; Sigma-Aldrich), Adenylyl cyclase III (C-20, IF: 1:100; Santa Cruz), B9D1 (NBP2-84489, WB: 1:1,000; Novus Biologicals) and HTR6 (31 (link)). Goat polyclonal antibodies: γ-Tubulin (sc-7396, IF: 1:200; Santa Cruz). Alexa Fluor (AF)–conjugated donkey secondary antibodies from Thermo Fisher Scientific (all used for IF at 1:10,000): AF488 anti-mouse IgG (A21202), AF488 anti-rabbit IgG (A21206), AF555 anti-mouse IgG (A31570), AF555 anti-rabbit IgG (A31572), AF594 anti-rabbit IgG (A21207), and AF647 anti-goat IgG (A21447). Also from Thermo Fisher Scientific were HRP-conjugated secondary antibodies (used for WB at 62 ng/ml): goat anti-rabbit IgG (A16104), goat anti-mouse IgG (A16072) and donkey anti-rat IgG (A18739). Biotinylated proteins were detected with Neutravidin-HRP (A2664, 1 μg/ml; Thermo Fisher Scientific). D-biotin was from Thermo Fisher Scientific (BP-232-1).

Barbeito P., Tachibana Y., Martin-Morales R., Moreno P., Mykytyn K., Kobayashi T, & Garcia-Gonzalo F.R. (2020). HTR6 and SSTR3 ciliary targeting relies on both IC3 loops and C-terminal tails. Life Science Alliance, 4(3), e202000746.

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Quantifying Protein Methylation Dynamics

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Doxorubicin and pemetrexed powder was purchased from Sigma Aldrich (Merck, Darmstadt, Germany) and diluted in dimethyl sulfoxide (DMSO). Stock solutions were stored at − 20 °C and diluted to the desired concentrations with growth medium before use. The PRMT5 (P4847), PRMT1 (G1544) and FLAG (F7425) antibodies were purchased from Sigma Aldrich (America). The CFLAR_L (ALX-804-961-0100) antibody was purchased from Enzo Biochem. The monomethyl arginine (8015S), CASP8 (9746 L) and PARP-1 (#9542) antibodies were purchased from Cell Signaling Technology (Boston, Massachusetts, US). The symmetric dimethyl arginine and asymmetric dimethyl arginine antibodies were purchased from Millipore.

Li M., An W., Xu L., Lin Y., Su L, & Liu X. (2019). The arginine methyltransferase PRMT5 and PRMT1 distinctly regulate the degradation of anti-apoptotic protein CFLARL in human lung cancer cells. Journal of Experimental & Clinical Cancer Research : CR, 38, 64.

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Cell Lines and Antibodies for Apoptosis

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The following cell lines were used: 293T (H. Young, Columbia University, College of Physicians and Surgeons, New York, NY), HeLa, and COS-7 (ATCC, Manassas, VA). They were maintained in DMEM supplemented with 10% heat-inactivated fetal calf serum (FCS). Suspension T lymphocyte cell lines MT1, MT2, MT4, and HuT102 were maintained in RPMI 1640 (Gibco), supplemented with 10% FCS. The CaGT cell line was established by culturing fresh peripheral blood mononuclear cells (PBMCs) of an ATL patient in a media consisting of 45% RPMI 1640 (Gibco), 45% HD SFM (Gibco), and 10% FCS without any exogenous cytokines or mitogens. Following 4 months in culture, clones were isolated by limiting dilution. Phenotypic analyses revealed a T-cell origin expressing CD2, CD4, CD3^dim, CD25, CD122, and CD132, and negative for CD8, CD7, CD19, and CD20. The following antibodies were used to detect epitope-tagged proteins: Flag (F-7425) (Sigma, St Louis, MO), HA (sc-805) (Santa Cruz Biotechnology, Santa Cruz, CA), and V5 (R960-25) (Invitrogen, Carlsbad, CA). Antibodies against caspase-3 (9662) (Cell Signaling Technologies, Beverly, MA), and PARP (556494) (BD Pharmingen, San Diego, CA) were used to detect apoptosis.

Nguyen V.P., Chen J., Petrus M.N., Goldman C.K., Kruhlak M.J., Bamford R.N, & Waldmann T.A. (2014). A New Domain in the Toll/IL-1R Domain-Containing Adaptor Inducing Interferon-ß Factor Protein Amino Terminus Is Important for Tumor Necrosis Factor-a Receptor-Associated Factor 3 Association, Protein Stabilization and Interferon Signaling. Journal of Innate Immunity, 6(3), 377-393.

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Comprehensive Antibody Panel for EMT Markers

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FBXL10 rabbit polyclonal antibody (09–864) was purchased from Merck Millipore. Rabbit monoclonal antibodies against SNAI1 (3879), E-cadherin (3195), N-cadherin (13116), Vimentin (5741) and acetylated-lysine (9441) were obtained from Cell Signaling Technology. Rabbit monoclonal antibody against Cytokeratin 18 (ab133272) was obtained from Abcam. GAPDH (sc-47724) and rabbit polyclonal p300 (sc-585) were purchased from Santa Cruz Biotechnology. Flag (F7425) and MYC (SAB5700727) antibodies were purchased from Sigma. GFP (GTX113617) and HA (GTX115044) were obtained from GeneTex. Protein A/G and Flag-affinity magnetic beads were purchased from Biomake. GFP-selector (2-9131-020) beads were obtained from IBA Life Sciences. MS-275 (S1503) was purchased from Selleck.

Yang Y., Zhao B., Lv L., Yang Y., Li S, & Wu H. (2021). FBXL10 promotes EMT and metastasis of breast cancer cells via regulating the acetylation and transcriptional activity of SNAI1. Cell Death Discovery, 7, 328.

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Evaluating Mitochondrial Function and Oxidative Stress

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Puromycin (P8833), Baf A1 (J61835), polybrene (H9268), irinotecan (I1406), all-trans-retinoic acid (R2625), and N-acetyl-l-cysteine (A9165) were all purchased from Sigma-Aldrich (USA). Anti-Top1mt–specific antibody (47 (link), 52 (link)) (Top1mt, polyclonal mouse) was a gift from Y. Pommier [Center for Cancer Research, National Cancer Institute, National Institutes of Health (NIH), USA]. Rabbit polyclonal antibodies [TDP1 (Ab4166), PGC1α (Ab54481), NRF1 (Ab34682), and mtTFAM (Ab47517)] and mouse monoclonal antibodies [COX IV (Ab14744), DRP1 (Ab56788), Mfn1 (Ab57602), and double-stranded DNA (dsDNA) (Ab27156)] were purchased from Abcam (Cambridge, USA). Mouse monoclonal antibodies [FLAG (M2) (F3165)] and rabbit polyclonal antibodies [FLAG (F7425) and LC3B (L7543)] were purchased from Sigma-Aldrich (St. Louis, MO, USA). Mouse monoclonal γH2AX (05-636) antibody was purchased from Millipore, USA. Rabbit monoclonal GFP (G10362) and anti OXPHOS complex kit (457999) antibodies were purchased from Invitrogen. Mouse monoclonal actin (SC-8432) and secondary antibodies [horseradish peroxidase–conjugated anti-rabbit immunoglobulin G (IgG) or anti-mouse IgG] were purchased from Santa Cruz Biotechnology (Santa Cruz, USA). MitoTracker red (M22426), MitoTracker green (M7514), TMRM (T668), CM-H₂DCFDA (C6827), and LysoTracker red DND-99 (L7528) were obtained from Molecular Probes.

Ghosh A., Bhattacharjee S., Chowdhuri S.P., Mallick A., Rehman I., Basu S, & Das B.B. (2019). SCAN1-TDP1 trapping on mitochondrial DNA promotes mitochondrial dysfunction and mitophagy. Science Advances, 5(11), eaax9778.

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Comprehensive Antibody Evaluation Protocol

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Anti‐acetyl‐lysine (Ac‐K; ab21623), anti‐Tip60 (ab137518), anti‐Myc (ab9106), anti‐p53 (ab26), anti‐p21 (ab188224), anti‐puma (ab9643), anti‐HA (ab18181) and anti‐KDM2B (ab137547) antibodies were purchased from Abcam (Cambridge, MA). Anti‐H3K36me2 (#2901) and anti‐H3K36me3 (#4909S) antibodies were purchased from Cell Signaling Technology (Danvers, MA). The antibody specific for Flag (F7425) was purchased from Sigma (St. Louis, MO).

Shi X, & Fan M. (2019). Tip60‐dependent acetylation of KDM2B promotes osteosarcoma carcinogenesis. Journal of Cellular and Molecular Medicine, 23(9), 6154-6163.

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Immunoblotting Reagents for Autophagy Proteins

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Primary antibodies against ATG13 (13468), ATG101 (13492), LC3B (2775) and WIPI2 (8567) were purchased from Cell Signaling Technology (Danvers, MA, USA); Antibodies against HUWE1 (A300-486), β-actin (A300–491A), HA (A190–108A) and WIPI2 (A305-324A) were purchased from Bethyl Laboratories(Montgomery, TX, USA); those against FLAG M2 (F1804) and FLAG (F7425) were purchased from Sigma Aldrich (St. Louis, MO, USA); antibody against GFP (mouse SC-9996)(rabbit SC 8334) were purchased from Santa Cruz Biotechnology; and antibody against p62(610832) was purchased from BD Bioscience; antibody against ATG14 (GTX119950) was purchased from GeneTex (Hsinchu, Taiwan). Secondary antibodies against horseradish peroxidase-linked anti-rabbit (A120–101P) and anti-mouse (A90–116P), were purchased from Bethyl Laboratories.
Hoechst 33342 (H3570), Lipofectamine^TM 2000 (11668019), Lipofectamine^TM RNAiMAX (13778150) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Rapamycin (Rapamycin from Streptomyces hygroscopicus, R0395), CCCP (carbonyl cyanide 3-chlorophenylhydrazone, C2759), Chloroquine (CQ, C6628), MG132 (M7449), and Cycloheximide (CHX, C4859) were purchased from Sigma Aldrich. Protease inhibitor cocktail tablets (11697498001) were purchased from Roche Applied Bioscience (Penzberg, Germany).

Lee J., Kim J., Shin J., Kang Y., Choi J, & Cheong H. (2021). ATG101 Degradation by HUWE1-Mediated Ubiquitination Impairs Autophagy and Reduces Survival in Cancer Cells. International Journal of Molecular Sciences, 22(17), 9182.

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