Liver tissues were homogenized and sonicated in RIPA lysis buffer (Sigma-Aldrich, St. Louis, MO) containing protease inhibitor cocktail and phosphostop (Roche Diagnostics, Indianapolis, IN), and total protein was extracted by centrifugation. A total of 30 μg protein was resolved on 4–12% precast SDS-PAGE gel (Invitrogen, Grand Island, NY) and transblotted onto a polyvinylidene fluoride membrane (Bio-Rad, Hercules, CA). Membranes were probed with antisera against p-JNK, JNK, p-PDK, PDK, p-AKT, AKT, p-PKCζ, PKCζ, p-PERK, PERK, p-elF2α, elF2α, p-IRE-1, IRE-1, GRP78, CHOP, caspase-3, cleaved caspase-3, ATF-6α, XBP-1, BCL-2, Bax, LC-3B and β-actin (Cell Signaling Technology, Danvers, MA). Isotype-matched horseradish peroxidase conjugated secondary antibodies, enhanced chemiluminescence substrate (Pierce, Rockford, IL) and a FluorChem 8900 digital imaging system (AlphaInnotech, San Leandro, CA) were used to visualize protein bands. Densitometric analyses was performed with VisionWorks® Software, version 6.8 (UVP, Upland, CA).
Fluorchem 8900 digital imaging system
Manufactured by Bio-Techne
The FluorChem 8900 digital imaging system is a high-performance device designed for capturing and analyzing fluorescent images. It features a cooled CCD camera, adjustable lighting, and specialized software for image acquisition and data analysis.
Automatically generated - may contain errors
Lab products found in correlation
Cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Precast sds page gel, by Thermo Fisher Scientific (2 mentions)
Polyvinylidene fluoride membrane, by Bio-Rad (2 mentions)
Ripa lysis buffer, by Merck Group (2 mentions)
Bcl2 associated x (bax), by Cell Signaling Technology (2 mentions)
P elf2α, by Cell Signaling Technology (2 mentions)
Caspase 3, by Cell Signaling Technology (2 mentions)
Bcl 2, by Cell Signaling Technology (2 mentions)
β actin, by Cell Signaling Technology (2 mentions)
Protease inhibitor cocktail and phosphostop, by Roche (2 mentions)
Grp78, by Cell Signaling Technology (2 mentions)
Elf2α, by Cell Signaling Technology (2 mentions)
P ire1, by Cell Signaling Technology (2 mentions)
Atf6α, by Cell Signaling Technology (2 mentions)
Visionworks software version 6, by Analytik Jena (2 mentions)
2 protocols using fluorchem 8900 digital imaging system
1
Western Blot Analysis of Liver Proteins
2
Western Blot Analysis of Liver Proteins
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Liver tissues were homogenized and sonicated in RIPA lysis buffer (Sigma-Aldrich, St. Louis, MO) containing protease inhibitor cocktail and phosphostop (Roche Diagnostics, Indianapolis, IN), and total protein was extracted by centrifugation. A total of 30 μg protein was resolved on 4–12% precast SDS-PAGE gel (Invitrogen, Grand Island, NY) and transblotted onto a polyvinylidene fluoride membrane (Bio-Rad, Hercules, CA). Membranes were probed with antisera against p-JNK, JNK, p-PDK, PDK, p-AKT, AKT, p-PKCζ, PKCζ, p-PERK, PERK, p-elF2α, elF2α, p-IRE-1, IRE-1, GRP78, CHOP, caspase-3, cleaved caspase-3, ATF-6α, XBP-1, BCL-2, Bax, LC-3B and β-actin (Cell Signaling Technology, Danvers, MA). Isotype-matched horseradish peroxidase conjugated secondary antibodies, enhanced chemiluminescence substrate (Pierce, Rockford, IL) and a FluorChem 8900 digital imaging system (AlphaInnotech, San Leandro, CA) were used to visualize protein bands. Densitometric analyses was performed with VisionWorks® Software, version 6.8 (UVP, Upland, CA).
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