Anti gst

Manufactured by Thermo Fisher Scientific

Sourced in United States

The Anti-GST product is a laboratory reagent used for the detection and purification of proteins fused with the Glutathione S-Transferase (GST) tag. It is designed to bind and capture GST-tagged proteins, facilitating their isolation and analysis.

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Lab products found in correlation

Anti ha, by Thermo Fisher Scientific (3 mentions) Anti flag, by Cell Signaling Technology (2 mentions) Anti tcf7l2, by Cell Signaling Technology (2 mentions) Anti ha, by Roche (2 mentions) Anti flag hrp, by Merck Group (2 mentions) Anti actin, by Merck Group (2 mentions) Anti sf3b1, by Fortis Life Sciences (2 mentions) Anti sugp1, by Fortis Life Sciences (2 mentions) Rabbit anti stat1, by Santa Cruz Biotechnology (1 mentions) Rabbit anti stat2, by Santa Cruz Biotechnology (1 mentions) Anti phospho stat2, by R&D Systems (1 mentions) Mouse anti gapdh, by Santa Cruz Biotechnology (1 mentions) Mouse anti v5, by Thermo Fisher Scientific (1 mentions) Anti phospho stat3 tyr705, by Cell Signaling Technology (1 mentions) Bca assay, by Thermo Fisher Scientific (1 mentions) Anti α tubulin, by Santa Cruz Biotechnology (1 mentions) Anti histone h3, by Abcam (1 mentions) Anti p tyr, by Santa Cruz Biotechnology (1 mentions) Anti flag, by Merck Group (1 mentions) Anti β actin, by Cell Signaling Technology (1 mentions)

Spelling variants of this product

LanthaScreen™ Eu anti-GST Antibody (11 citations) Alexa488-conjugated anti-GST antibody (5 citations) Mouse anti-GST (3 citations) Lantha-screen Tb anti-GST antibody (3 citations) Anti-GST antibody-coated microtiter plates (2 citations) Tb anti-GST antibody (2 citations) Eu-anti-GST antibody (2 citations) Alexa Fluor 488-conjugated anti-GST antibody (2 citations) LanthaScreen Eu‐anti‐GST (2 citations) Anti-GST tag polyclonal Ab (2 citations)

18 protocols using anti gst

Characterization of JAK-STAT Signaling

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Dual luciferase assay, confocal immunofluorescence microscopy, immunoprecipitation and Western blotting were performed as previously described [50 (link)–52 (link)]. Relative luciferase activity in arbitrary units was calculated by normalizing firefly luciferase activity with Renilla luciferase activity.
Mouse anti-c-myc (clone 9E10) was purchased from MilliporeSigma. Mouse anti-V5 and anti-GST were from Thermo Fisher Scientific. Rabbit anti-STAT1, rabbit anti-STAT-2, mouse anti-JAK1 (A-9), and mouse anti-GAPDH were purchased from Santa Cruz Biotechnology (TX, USA). Rabbit polyclonal antibodies against phospho-STAT1, phospho-JAK1 were bought from Cell Signaling Technology (MA, USA). Anti-phospho-STAT2 was purchased from R&D Systems (MN, USA).

Fung S.Y., Siu K.L., Lin H., Chan C.P., Yeung M.L, & Jin D.Y. (2022). SARS-CoV-2 NSP13 helicase suppresses interferon signaling by perturbing JAK1 phosphorylation of STAT1. Cell & Bioscience, 12, 36.

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Western Blot Analysis of Protein Signaling

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Cells were lysed using MAPK lysis buffer (50 mM HEPES, 4 mM sodium pyrophosphate, 10 mM sodium fluoride, 2 mM orthovanadate, 100 mM NaCl, 10 mM EDTA, pH 7.5) with protease inhibitor cocktail (Sigma). Protein concentrations were determined by BCA assay (Thermo Scientific). Proteins were separated by SDS-PAGE and transferred to nitrocellulose membranes. After blocking with non-fat milk, the membrane was probed with primary antibodies against proteins of interest. The primary antibodies included anti-STAT3pan (Cell Signaling Technology 124H6), anti-phospho-STAT3 (Tyr705) (Cell Signaling Technology #9145), anti-JAK1 (Millipore/Upstate 06-272), anti-GST (Thermo Scientific MA4-004), anti-Flag (Sigma F1804), anti-HA (Thermo Scientific 26183), anti-pTyr (Santa Cruz Biotechnology sc-18182), anti-α-tubulin (Santa Cruz Biotechnology sc-8035), anti-β-actin (Cell Signaling Technology #4967), and anti-histone-H3 (Abcam#ab1791). After blotting with the HRP-conjugated secondary antibody, the membrane was developed using a chemiluminescence HRP substrate kit (Thermo Scientific).

Zhu F., Hwang B., Miyamoto S, & Rui L. (2016). Nuclear Import of JAK1 is Mediated by a Classical NLS and is Required for Survival of Diffuse Large B-cell Lymphoma. Molecular cancer research : MCR, 15(3), 348-357.

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Protein Binding Microarray Analysis Protocol

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We employed an “all 10-mer” universal array design in 8 × 3 60K, GSE format (Agilent Technologies; AMADID #030236). Double-stranding of oligonucleotide arrays and PBM experiments were performed following previously described experimental protocols (Berger and Bulyk, 2006 (link), 2009 (link); Berger et al., 2006 (link)). Briefly, the TF of interest was expressed with an epitope tag (GST, HA, or FLAG), applied to the double-stranded DNA array, and detected with fluorescently labeled antibody specific for the tag. Experimental conditions used for all PBM experiments, including TF concentrations and buffers, are provided in Table S5. The following Alexa Fluor 488-conjugated antibodies were used: anti-GST (Thermo Fisher Scientific, A-11131), anti-HA (Thermo Fisher Scientific, A-21287), and anti-FLAG (Cell Signaling, 5407).

Shokri L., Inukai S., Hafner A., Weinand K., Hens K., Vedenko A., Gisselbrecht S.S., Dainese R., Bischof J., Furger E., Feuz J.D., Basler K., Deplancke B, & Bulyk M.L. (2019). A Comprehensive Drosophila melanogaster Transcription Factor Interactome. Cell reports, 27(3), 955-970.e7.

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Triad Vesicles Containing TRPC3

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The triad vesicles containing TRPC3 were prepared from rabbit fast-twitch skeletal muscle and were solubilized, as previously described [4 (link),6 (link),25 (link),29 (link)]. All surgical interventions and methods of animal care were conducted under the guidelines, as previously described [6 (link),27 (link),30 (link)]. The solubilized triad sample or the lysate of myotubes was subjected to co-immunoprecipitation assay using anti-TRPC3 antibody [28 (link),30 (link)]. For immunoblot assays, various antibodies were used: anti-RyR1, anti-DHPR, anti-SERCA1a, anti-MG29, anti-JP1, anti-JP2, and anti-GST antibodies (1:1,000) from Thermo Scientific Inc. (Rockford, IL, USA), anti-TRPC3 and anti-TRPC4 antibodies (1:800) from Alomone Laboratories (Jerusalem 9104201, Israel), and anti-Orai1, anti-STIM1, and anti-α-actin antibodies (1:1,000) from Abcam (Cambridge, MA, USA).

Woo J.S., Hwang J.H., Huang M., Ahn M.K., Cho C.H., Ma J, & Lee E.H. (2015). Interaction between mitsugumin 29 and TRPC3 participates in regulating Ca2+ transients in skeletal muscle. Biochemical and biophysical research communications, 464(1), 133-139.

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Protein Binding Microarray Analysis Protocol

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Recombinant BP180 Protein Immunoblotting

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FL-BP180 was expressed in COS7 cells transfected with human BP180 cDNA (35 (link)) and prepared as described previously (36 (link)). Fifty ng of each recombinant human glutathione-S-transferase (GST)-BP180-fusion protein expressed in E. coli spanning most of the BP180 polypeptide were used as an antigen. Immunoblotting and preparation of GST-BP180 fusion proteins were performed as described previously (30 (link)). Serum samples were diluted to 1:100 in 5% non-fat milk-TBS-0.1% Tween-20 and 1:50 000 peroxidase-conjugated anti-human IgG (Sigma-Aldrich, St. Louis, MO, USA) was used as a secondary antibody. Anti-GST (1:3000, Thermo Fisher Scientific, Rockford, IL, USA) with peroxidase-conjugated anti-rabbit IgG (Sigma-Aldrich) were used to detect fusion proteins. Protein bands were visualized with ECL Prime substrate (GE Healthcare, Buckinghamshire, UK) on a LAS Imager 3000 (Fujifilm, Tokyo, Japan). Epitope mapping data of 14 age and sex-matched healthy controls from our previous work (30 (link)) were used as a control.

Nätynki A., Leisti P., Tuusa J., Varpuluoma O., Huilaja L., Izumi K., Herukka S.K., Ukkola O., Junttila J., Kokkonen N, & Tasanen K. (2022). Use of gliptins reduces levels of SDF-1/CXCL12 in bullous pemphigoid and type 2 diabetes, but does not increase autoantibodies against BP180 in diabetic patients. Frontiers in Immunology, 13, 942131.

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Affinity Purification of HaloTag Proteins

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BL21(DE3) expressing HaloTag or HaloTag-PLEKHN1 were lysed in PBS with 1 mM βME and 1 mM MgCl₂ (supplemented with 1 mM PMSF and 0.5 mM benzamidine) using a LV1 Microfluidics and spun at 24,000×g for 35 min. Of this lysate, 500 μL were added to MagneHaloTag beads (Promega, Madison, WI) pre-equilibrated with wash buffer (PBS with 0.005% NP-40) and incubated overnight at 4 °C. The lysate was then removed and the beads were washed with wash buffer three times. 500 μL of lysate from BL21(DE3) expressing GST-AnkX prepared as indicated above was added to the HaloTag and HaloTag-PLEKHN1 coated beads and incubated for 2 h at 4 °C. The lysate was then removed and the beads were washed with wash buffer three times. The bound proteins were eluted via boiling in Laemmli buffer. The lysates and eluates were separated via SDS-PAGE and transferred to a PVDF membrane for immunoblot analysis. Fast western kit was used in conjunction with primary antibodies: anti-HaloTag (Promega) at 1:1000 and anti-GST (Thermo Scientific, MA4–004) at 1:1000.

Yu X., Noll R.R., Romero Dueñas B.P., Allgood S.C., Barker K., Caplan J.L., Machner M.P., LaBaer J., Qiu J, & Neunuebel M.R. (2018). Legionella effector AnkX interacts with host nuclear protein PLEKHN1. BMC Microbiology, 18, 5.

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Western Blotting of Protein Targets

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The liver tissues and cell lines were lysed using radioimmunoprecipitation assay (RIPA) buffer (#P1003B, Beyotime). Approximately 30 μg of total protein was added to 8% SDS PAGE gels using Bio-Rad equipment (Shanghai, China). After separation, the proteins were transferred to PVDF membranes (#ISEQ00010, Millipore, Massachusetts, USA). The protein-loaded PVDF membranes were blocked with 5% skim milk (#A600669, Sangon), immersed in solutions of the primary antibodies at 4 °C overnight, and then incubated with Alexa Fluor 680-conjugated secondary antibodies (Thermo Fisher Scientific). Tubulin-α or GAPDH was used as an internal control. The used primary antibodies were listed (anti-NLRP3, #IMG-6668A, Novus Biologicals, Colorado, USA; anti-MARCH7, #PA5-54572, Sigma Aldrich; anti-Cleaved caspase-1, #PA5-99390, Thermo Fisher Scientific; anti-Matured IL-1β, #AF401, R&D Systems, City of Emeryville, USA; anti-GSDMD-N, #GSDMD antibody Abcam EPR 19828, Abcam, Cambridge, UK; anti-Tubulin-α, #NB100-690, Novus Biologicals; anti-GAPDH, #MA1-16757, Thermo Fisher Scientific; anti-actin-β, #NB600-501, Novus Biologicals; anti-HA, #NB600-363, Novus Biologicals; anti-Flag, #MA1-91878, Thermo Fisher Scientific; anti-Ubiquitin, #NB300-130, Novus Biologicals; anti-GST, #13-6700, Thermo Fisher Scientific; anti-His, #NBP2-61482, Novus Biologicals; anti-Myc, #2276, Cell Signaling Technology, Boston, USA).

Chen T., Meng Y., Zhou Z., Li H., Wan L., Kang A., Guo W., Ren K., Song X., Chen Y, & Zhao W. (2023). GAS5 protects against nonalcoholic fatty liver disease via miR-28a-5p/MARCH7/NLRP3 axis-mediated pyroptosis. Cell death and differentiation, 30(7).

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GluA1 C-terminus Phosphorylation Assay

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Purified GST fusion proteins (20 μl) with a GluA1 C terminus were subjected to an in vitro phosphorylation assay using the CAMK2a Kinase Enzyme System and PKA Kinase Enzyme System according to the manufacturer’s protocol (Promega). Phosphorylated GST fusion proteins were analyzed by immunoblot analysis using anti-Phospho-GluA1 (Ser831), Phospho-GluA1 (Ser845) (Invitrogen), and anti-GST (Amersham) antibodies.

Matsuda S, & Yuzaki M. (2021). Subunit-dependent and subunit-independent rules of AMPA receptor trafficking during chemical long-term depression in hippocampal neurons. The Journal of Biological Chemistry, 297(2), 100949.

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GST-MinD Interaction with His-cyto-AtoS

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GST-MinD (5 μM) was incubated with His-cyto-AtoS (5 μM) in buffer-B (50 mM Hepes pH 7.4, 150 mM KCl) at room temperature for 30 min. HisPure cobalt resin was added to the reaction mixture and incubated for 1 h at 4 °C. His-cyto-AtoS (5 μM) or GST-MinD (5 μM) alone were used as controls. The whole reaction mixture was transferred to a spin column, washed three times with buffer B containing 20 mM imidazole and eluted with buffer containing 50 mM Hepes pH 7.4, 150 mM KCl, and 300 mM imidazole. The GST-tagged MinD fraction eluted along with His-cyto-AtoS was verified on a 12% SDS-PAGE and also subjected to Western blot analysis. Anti-GST (Invitrogen: 136700), anti-His primary antibodies (Sigma), and anti-mouse-horseradish peroxidase–conjugated secondary antibodies (Sigma) were used for the detection of MinD and cyto-AtoS. All antibodies were used at a 1:10,000 dilution.

Pradhan P., Taviti A.C, & Beuria T.K. (2024). The bacterial division protein MinDE has an independent function in flagellation. The Journal of Biological Chemistry, 300(4), 107117.

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