Eosinophil or neutrophil cell pellets (from 15 million cells) were lysed in buffer containing 10mM Tris HCl (pH 7.48), 0.1 mM NaCl, 1mM EDTA, 1mM EGTA, 1mM NaF, 14mM Na4P2O7, 2mM Na3VO4, 0.1% Triton X-100, 0.1% SDS, mammalian cocktail protease inhibitors (Sigma-Aldrich Corp, St. Louis, MO, USA) and 1mM PMSF.46 (link) The suspension was sonicated five times with 5 second pulses (output setting 0.5, Sonicator 3000, Misonix, Farmingdale, NY, USA), repeatedly passed through a syringe (28 gauge needle), and clarified by centrifugation (12 000 x g/10 min/4 °C). The lysate was incubated at 4°C for 24–30 h with protein G-coupled magnetic beads (EMD Millipore Corp, Billerica, MA, USA) coated with polyclonal goat anti-human activin A antibody (R&D Systems). The beads were collected with a magnet and resuspended in 20 μl of non-reducing lithium dodecyl sulfate sample buffer (Thermo Fisher Scientific, Grand Island, NY, USA). Samples were heated to 90°C for 5 min and loaded onto a 13.5 % SDS-PAGE gel.
Protein g coupled magnetic beads
Manufactured by Merck Group
Sourced in United States
Protein G-coupled magnetic beads are a type of laboratory equipment used in various scientific applications. They consist of magnetic particles coated with protein G, a bacterial cell surface protein that has a high affinity for the Fc region of immunoglobulins. These beads can be used to capture and separate antibodies or other proteins from complex biological samples.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using protein g coupled magnetic beads
1
Activin A Immunoprecipitation from Leukocytes
2
Activin A Immunoprecipitation from Leukocytes
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Eosinophil or neutrophil cell pellets (from 15 million cells) were lysed in buffer containing 10mM Tris HCl (pH 7.48), 0.1 mM NaCl, 1mM EDTA, 1mM EGTA, 1mM NaF, 14mM Na4P2O7, 2mM Na3VO4, 0.1% Triton X-100, 0.1% SDS, mammalian cocktail protease inhibitors (Sigma-Aldrich Corp, St. Louis, MO, USA) and 1mM PMSF.46 (link) The suspension was sonicated five times with 5 second pulses (output setting 0.5, Sonicator 3000, Misonix, Farmingdale, NY, USA), repeatedly passed through a syringe (28 gauge needle), and clarified by centrifugation (12 000 x g/10 min/4 °C). The lysate was incubated at 4°C for 24–30 h with protein G-coupled magnetic beads (EMD Millipore Corp, Billerica, MA, USA) coated with polyclonal goat anti-human activin A antibody (R&D Systems). The beads were collected with a magnet and resuspended in 20 μl of non-reducing lithium dodecyl sulfate sample buffer (Thermo Fisher Scientific, Grand Island, NY, USA). Samples were heated to 90°C for 5 min and loaded onto a 13.5 % SDS-PAGE gel.
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