Exosome and cell pellets were lysed in sodium dodecyl sulfate (SDS)-urea lysis buffer [5% SDS, 9 M urea, 125 mMTris HCI pH 6.8 supplemented with protease inhibitor cocktail (Sigma P8340)]. Protein concentration was determined by bicinchoninic acid (BCA, Pierce, Thermo Scientific) and 10 μg were dissolved into SDS loading buffer (with 5% βME) and fractionated on 4–15% Mini-protean TGX gels (BioRad, Hercules CA, USA) in 1x Tris Glycine SDS (TGS) 1× running buffer at room temperature. Proteins were electro-transferred to PVDF (Immobilon-P, Millipore) in 1x TGS buffer-methanol (20%) buffer. The blots were blocked with 5% milk Tween-20 (0.1%) PBS buffer (T-PBS, pH 7.4) for 1 h and incubated overnight at 4°C in primary antibodies. Antibodies were: anti-Alix/PDC61 (Ab76608) and anti-GRP4 (Cell Signaling #2104). Secondary antibodies were HRP-conjugated anti-rabbit IgG and anti-mouse IgG from Cell Signaling and Southern Biotech (Birmingham, AL, USA), respectively.
Anti alix pdc61
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-Alix/PDC61 is a primary antibody that recognizes the Alix/PDC6I protein. Alix/PDC6I is a cytoplasmic protein involved in diverse cellular processes, including cell division, endocytosis, and apoptosis. This antibody can be used to detect and study the Alix/PDC6I protein in various experimental applications.
Automatically generated - may contain errors
Lab products found in correlation
Anti grp4, by Cell Signaling Technology (2 mentions)
Hrp conjugated anti rabbit igg, by Cell Signaling Technology (2 mentions)
Immobilon p, by Merck Group (2 mentions)
Protease inhibitor cocktail, by Merck Group (2 mentions)
Mini protean tgx gel, by Bio-Rad (2 mentions)
Anti β actin, by Merck Group (1 mentions)
Anti hsp90, by Cell Signaling Technology (1 mentions)
Anti cd63, by Abcam (1 mentions)
2 protocols using anti alix pdc61
1
Exosome Protein Extraction and Analysis
2
Extracellular Vesicle Protein Analysis
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Exosome and cell pellets were lysed in sodium dodecyl sulfate (SDS)-urea lysis buffer [5% SDS, 9 M urea, 125 mM Tris HCl pH 6.8 supplemented with protease inhibitor cocktail (Sigma P8340)]. Protein concentration was determined by bicinchoninic acid (BCA, Pierce, Thermo Scientific) and 10 μg were dissolved into SDS loading buffer (with 5% βME) and fractionated on 4–15% Mini-protean TGX gels (BioRad, Hercules CA, USA) in 1× Tris Glycine SDS (TGS) 1× running buffer at room temperature. Proteins were electro-transferred to PVDF (Immobilon-P, Millipore) in 1× TGS buffer-methanol (20%) buffer. The blots were blocked with 5% milk Tween-20 (0.1%) PBS buffer (T-PBS, pH 7.4) for 1 h and incubated overnight at 4°C in primary antibodies. Antibodies were: anti-CD63 (Abcam ab193349); anti-HSP90 (Cell Signaling C45G5, Danvers, MA, USA); anti-Alix/PDC61 (Ab76608); Anti-GRP4 (Cell Signaling #2104); and anti- β-actin (Sigma, AC-15). Secondary antibodies were HRP-conjugated anti-rabbit IgG and anti-mouse IgG from Cell Signaling and Southern Biotech (Birmingham, AL, USA), respectively.
Public submission : We have submitted all relevant data of our experiments to the EV-TRACK knowledgebase (EV-TRACK ID: EV170017).[37 (link)]
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