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Mouse monoclonal anti gst antibody

Manufactured by Thermo Fisher Scientific

The mouse monoclonal anti-GST antibody is a laboratory reagent used for the detection and quantification of glutathione S-transferase (GST) in various assays and applications.

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2 protocols using mouse monoclonal anti gst antibody

1

Visualizing Recombinant and Patient Derived Poly(GA) Proteins

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To examine the filamentous structure of recombinant GST-(GA)50 proteins, recombinant GST or GST-(GA)50 proteins were diluted to 1 µg/µl in 20 mM Tris–HCl, pH 7.5, 50 mM KCl, 10 mM MgCl2 in a final volume of 30 µl. The samples were incubated at 30 °C for 6 h, and then diluted to 0.1 µg/µl by reaction buffer and loaded onto grids for regular EM analysis. For immuno-EM analysis, mouse monoclonal anti-GST antibody (1:20, Thermo Scientific) was used as primary antibody, and goat anti-mouse IgG conjugated with 6 nm colloidal gold particles (1:20, Jackson ImmunoResearch Laboratories) was used as the secondary antibody. To examine the filamentous structure of poly(GA) proteins in c9FTD/ALS patients, small pieces (1.5 × 1.5 × 1 mm) of cerebellar folia or hippocampus from formalin-fixed brains were dissected and processed for routine electron microscopy (EM) or post-embedding immunogold EM as previously described [34 (link)]. Rabbit polyclonal anti-poly(GA) antibody (1:50) was used as a primary antibody and goat anti-rabbit IgG conjugated with 18 nm colloidal gold particles (1:20, Jackson ImmunoResearch Laboratories) was used as the secondary antibody. Thin sections stained with uranyl acetate and lead citrate were examined with a Philips 208S electron microscope (FEI) fitted with a Gatan 831 Orius CCD camera (Gatan). Digital images were processed with Adobe Photoshop CS5 software.
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2

GST-FolE Protein Overexpression and Purification

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Protein levels were determined by SDS-PAGE and Coomassie blue staining. Samples of the overexpression and purification of the GST-FolE protein in SDS sample buffer (750 mM Tris-HCl, pH 6.8, 40% glycerol, 0.002% Bromophenol blue, 8% SDS, 5% β-ME) were electrophoresed across a 12% Criterion TGX polyacrylamide gel (Bio-Rad, Hercules, CA) in running buffer (25 mM Tris, pH 8.3, 192 mM glycine, 0.1% SDS) at 200 V for 55 minutes. Proteins were stained with Coomassie Brilliant Blue R-250 (Fisher Scientific) and documented using an AlphaImager HP (Alpha Innotech, San Leandro, CA) with transilluminating white light. The overexpression of GST-FolE was verified by Western blot using a PVDF (polyvinyl difluoride) membrane (Bio-Rad, Hercules, CA) and probing with a 1:500 dilution of mouse monoclonal anti-GST antibody (Thermo Scientific, Waltham, MA) for 2 hours at room temperature. The membrane was then incubated with a 1:20,000 dilution of Immun-Star Goat Anti-Mouse-Horse Radish Peroxidase (HRP)-conjugated secondary antibody (Bio-Rad, Hercules, CA) for 1 hour at room temperature followed by documentation using Clarity Western ECL Substrates (Bio-Rad, Hercules, CA) and the C-Digit digital imager (Li-Cor, Lincoln, NE).
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