For real-time quantitative PCR (qPCR), brain tissues were homogenized in Trizol and total RNA was extracted using the Aurum Total RNA kit for fatty and fibrous tissue (Bio-Rad, Hercules, CA, USA). qPCR were performed as previously described (Martins-Silva et al., 2011 (link)). Primer sequences: STIP1-F, 5′-GCCAAGAAAGGAGACTACCAG-3′; STIP1-R, 5′-TCATAGGTTCGTTTGGCTTCC-3′; HsP90-F, 5′-CCACCCTGCTCTGTACTACT-3′; HsP90-R, 5′-CCAGGGCATCTGAAGCATTA-3′; HsP70-R, 5′-ACCTTGACAGTAATCGGTGC-3′; HsP70-F, 5′-CTCCCGGTGTGGTCTAGAAA-3′; PRP-F, 5′-GAACCATTTCAACCGAGCTG-3′; PRP-R, 5′-CATAGTCACAAAGAGGGCCAG-3′; Actin-F, 5′-TGGAATCCTGTGGCATCCATGA-3′; and Actin-R, 5′-AATGCCTGGGTACATGGTGGTA-3′. Immunoblot analysis was carried out as described previously (Beraldo et al., 2013 (link)). The antibodies used were anti-STIP1 (1:5000, in-house antibody generated by Bethyl Laboratories Montgomery, USA using recombinant STIP1) (Beraldo et al., 2013 (link)), anti-Hsp90 (1:1000), anti-Hsp70 (1:1000), anti-Hsp90α (1:1000), anti Hsp90β (1:1000) (Cell Signaling, Danvers, USA) and anti-PrP 8H4 (1:2000) (Abcam, Cambrige, UK).
Anti hsp90α
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-HSP90α is a laboratory reagent used to detect and quantify the HSP90α protein in biological samples. HSP90α is a molecular chaperone that plays a crucial role in the folding, stability, and function of various client proteins. The Anti-HSP90α product provides a specific and reliable tool for researchers to investigate the expression and activity of HSP90α in their experimental systems.
Automatically generated - may contain errors
Lab products found in correlation
Anti hsp90β, by Cell Signaling Technology (4 mentions)
Ripa lysis buffer, by Thermo Fisher Scientific (4 mentions)
Anti hsp90, by Cell Signaling Technology (2 mentions)
Anti ire1α, by Cell Signaling Technology (2 mentions)
Anti atf6, by Abcam (2 mentions)
Bicinchoninic acid assay, by Beyotime (2 mentions)
Anti β actin, by Santa Cruz Biotechnology (2 mentions)
Anti grp78, by Cell Signaling Technology (2 mentions)
Anti caspase 12 ab62484, by Abcam (2 mentions)
Anti p ire1α, by Thermo Fisher Scientific (2 mentions)
Anti grp78, by Abcam (2 mentions)
Anti hsp70, by Cell Signaling Technology (1 mentions)
Rabbit anti actin, by Cell Signaling Technology (1 mentions)
4 protocols using anti hsp90α
1
Quantitative PCR and Immunoblot Analysis of Brain Protein Expression
2
Western Blot Analysis of ER Stress Markers
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The protocol of Western blotting was similar with our previous study (65 (link)). The brain tissue or cell sample was sonicated in radioimmunoprecipitation assay (RIPA) lysis buffer (Thermo Fisher Scientific, Rockford, IL, USA) containing protease and phosphatase inhibitors. Samples were centrifuged at 12,000g for 20 min at 4°C, and the protein concentrations were examined by the bicinchoninic acid assay (Beyotime Biotechnology, Haimen, China). All the protein samples were heated for 5 min at 95°C in loading buffer. The deactivated protein samples were separated by SDS–polyacrylamide gel electrophoresis and then transferred to nitrocellulose membranes. The transferred membranes were incubated with primary antibodies and secondary antibodies. Anti-ATF6 (#ab37149; 1:500), anti–caspase 12 (ab62484; 1:1000), and anti-GRP78 (#ab21685; 1:1000) were purchased from Abcam (Cambridge, MA, USA). Anti-IRE1α (#3294; 1:500), anti-GRP78 (#3183; 1:1000), anti-HSP90α (#8165; 1:1000), and anti-HSP90β (#7411; 1:1000) were obtained from Cell Signaling Technology (San Francisco, CA, USA). Anti–p-IRE1α (#PA1-16927; 1:500) was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Anti–β-actin (#sc-47778; 1:2000) was obtained from Santa Cruz Biotechnology (Waltham, MA, USA).
3
Antibody Sourcing for PRRSV Research
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GA and 17-AAG obtained from Invivogen (San Diego, CA, USA) were re-suspended in DMSO.
Rabbit anti-actin, anti-HSP90α, anti-HSP90β, anti-HSP90 antibodies were obtained from Cell Signaling Technology (Beverly, MA, USA). Mouse anti-PRRSV N protein antibody was obtained from Jeno Biotech Inc (Chuncheon, South Korea). Mouse monoclonal antibody specific for dsRNA (J2) was purchased from Scicons (Hungary).
Rabbit anti-actin, anti-HSP90α, anti-HSP90β, anti-HSP90 antibodies were obtained from Cell Signaling Technology (Beverly, MA, USA). Mouse anti-PRRSV N protein antibody was obtained from Jeno Biotech Inc (Chuncheon, South Korea). Mouse monoclonal antibody specific for dsRNA (J2) was purchased from Scicons (Hungary).
4
Western Blot Analysis of ER Stress Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The protocol of Western blotting was similar with our previous study (65 (link)). The brain tissue or cell sample was sonicated in radioimmunoprecipitation assay (RIPA) lysis buffer (Thermo Fisher Scientific, Rockford, IL, USA) containing protease and phosphatase inhibitors. Samples were centrifuged at 12,000g for 20 min at 4°C, and the protein concentrations were examined by the bicinchoninic acid assay (Beyotime Biotechnology, Haimen, China). All the protein samples were heated for 5 min at 95°C in loading buffer. The deactivated protein samples were separated by SDS–polyacrylamide gel electrophoresis and then transferred to nitrocellulose membranes. The transferred membranes were incubated with primary antibodies and secondary antibodies. Anti-ATF6 (#ab37149; 1:500), anti–caspase 12 (ab62484; 1:1000), and anti-GRP78 (#ab21685; 1:1000) were purchased from Abcam (Cambridge, MA, USA). Anti-IRE1α (#3294; 1:500), anti-GRP78 (#3183; 1:1000), anti-HSP90α (#8165; 1:1000), and anti-HSP90β (#7411; 1:1000) were obtained from Cell Signaling Technology (San Francisco, CA, USA). Anti–p-IRE1α (#PA1-16927; 1:500) was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Anti–β-actin (#sc-47778; 1:2000) was obtained from Santa Cruz Biotechnology (Waltham, MA, USA).
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