Proteins were extracted from the cortex, hippocampus and cerebellum as described above (see Sample preparation for LC-MS/MS analysis section). 20 μg of proteins from each tissue lysate were resolved by 4–20% Mini-PROTEAN TGX precast gels (BioRad) and transferred to a PVDF membrane (BioRad). The membrane was first blocked in the blocking buffer (5% nonfat dry milk, 0.1% Tween-20; in PBS) for 1 h at room temperature and then incubated with anti-Aβ antibody (1:500; 4G8; Signet, USA), anti-HSP70 antibody (1:200; Santa Cruz Biotechnology, USA), anti-mTOR antibody (1:250; Santa Cruz Biotechnology, USA), and anti-Cdk-5 antibody (1:200; Santa Cruz Biotechnology, USA) in blocking buffer for overnight at 4°C. After washing, the membrane was incubated with an HRP-conjugated goat anti-mouse secondary antibody (1:2000; Santa Cruz Biotechnology, USA) for 1 h at room temperature and detected by chemiluminescence (Bio-Rad). Equal protein loading was confirmed by stripping the blot and re-probing with a β-actin antibody (1:1000; Santa Cruz Biotechnology, USA). Western blots were quantified by densitometry using the ImageJ software.
Anti hsp70 antibody
Manufactured by Santa Cruz Biotechnology
Sourced in United States
The Anti-HSP70 antibody is a primary antibody that specifically binds to the Heat Shock Protein 70 (HSP70) antigen. HSP70 is a highly conserved protein that plays a crucial role in protein folding and transport within cells. This antibody can be used to detect and quantify the expression levels of HSP70 in a variety of sample types.
Automatically generated - may contain errors
Lab products found in correlation
Clarity max ecl substrate, by Bio-Rad (2 mentions)
Anti cd81 antibody, by Abcam (2 mentions)
Anti calnexin antibody, by Santa Cruz Biotechnology (2 mentions)
Chemidoc mp gel imaging system, by Bio-Rad (2 mentions)
Anti tsg101 antibody, by Santa Cruz Biotechnology (2 mentions)
β actin antibody, by Santa Cruz Biotechnology (1 mentions)
Hrp conjugated goat anti mouse secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Anti mtor antibody, by Santa Cruz Biotechnology (1 mentions)
Pvdf membrane, by Bio-Rad (1 mentions)
Mini protean tgx precast gel, by Bio-Rad (1 mentions)
4 protocols using anti hsp70 antibody
1
Protein Expression Analysis in Alzheimer's
2
Isolation and Characterization of HSP70
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4T1 or CT26 cancer tissue was dissociated through mechanical dissection and utilized to prepare a single-cell suspension with the 70 μm cell strainer. Red blood cell lysis buffer was added to discard red blood cells. The prepared cancer cell was then homogenized on ice for 1 h in lysis buffer including Protease Inhibitor Cocktail Set I. Following centrifugation at 4 °C for 30 min, the supernatant was immunoprecipitated using anti-HSP70 antibody (Santa Cruz, sc-24) and incubated with protein A/G-Sepharose beads for 12 h. The lysis buffer was then used to wash the protein/antibody/beads complex. The complex was incubated with ATP at 25 °C for 30 min. Afterwards, the supernatant was gathered and quantitative measurement of protein was performed with Bradford assay. The protein was fractionated by SDS-PAGE electrophoresis and the gel was stained with Coomassie blue. The diameter of isolated HCP was measured using dynamic light scattering (DLS, Malvern, U.K.) with Zetasizer Software 7.01.
3
Western Blot Analysis of Exosomal Markers
4
Western Blot Analysis of Exosomal Markers
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Western blots were performed following SDS PAGE using standard protocols involving anti-CD81 antibody (Cat# ab232390) from Abcam, Waltham, MA, USA; anti-TSG101 antibody (Cat# SC-7964), anti-HSP70 antibody (Cat# SC-24) and anti-Calnexin antibody (Cat# SC-23954) from Santa Cruz Biotechnology, CA, USA. Following incubation with appropriate secondary antibody, proteins were detected by chemiluminescence using Clarity Max ECL substrate (Bio-Rad Laboratories, Hercules, California, USA) on a Chemidoc-MP Gel imaging system (Bio-Rad Laboratories, Hercules, California, USA).
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