Total protein extraction and mitochondrial and cytosolic fractions were analyzed by western blotting. Antibodies used for western blotting included antibodies that recognize PUMA, GRP78, sXBP-1 (Novus Biologicals, Littleton, CO, USA), eIF2α, p-eIF2α, cleaved caspase-12, cleaved caspase-4, cleaved caspase-9, cleaved caspase-3, Cox IV (all from Cell Signaling Technology), cytochrome c (Santa Cruz), Bax, Bak (Abcam) and β-actin (Sigma). Appropriate horseradish peroxidase-conjugated secondary antibodies were used to detect the primary antibody/antigen complexes. The signal was detected using ECL Western blotting detection reagents (Amersham Pharmacia Biotech, Piscataway, NJ, USA). After quantifying the signals by densitometry, the results are expressed as a ratio to loading control densitometry units.
Cleaved caspase 4
Manufactured by Cell Signaling Technology
Sourced in United States, Switzerland
Cleaved caspase-4 is an antibody that detects the active, cleaved form of caspase-4 protein. Caspase-4 is a member of the caspase family of proteases and is involved in the process of apoptosis, or programmed cell death.
Automatically generated - may contain errors
Lab products found in correlation
Cleaved caspase 3, by Cell Signaling Technology (2 mentions)
Cox 4, by Cell Signaling Technology (2 mentions)
Cleaved caspase 9, by Cell Signaling Technology (2 mentions)
Cleaved caspase 12, by Cell Signaling Technology (2 mentions)
Ecl western blotting detection reagent, by Cytiva (1 mentions)
Bcl2 associated x (bax), by Abcam (1 mentions)
β actin, by Merck Group (1 mentions)
P eif2α, by Cell Signaling Technology (1 mentions)
Cytochrome c, by Santa Cruz Biotechnology (1 mentions)
Goat anti mouse hrp, by Santa Cruz Biotechnology (1 mentions)
Goat anti rabbit hrp, by Santa Cruz Biotechnology (1 mentions)
Chicken anti rabbit alexa 594, by Thermo Fisher Scientific (1 mentions)
Donkey anti goat hrp, by Santa Cruz Biotechnology (1 mentions)
Mmp 7, by R&D Systems (1 mentions)
Chicken anti rabbit ap, by Santa Cruz Biotechnology (1 mentions)
Grp78, by Enzo Life Sciences (1 mentions)
Nuclear fast red, by Vector Laboratories (1 mentions)
Grp78, by Cell Signaling Technology (1 mentions)
Grp94, by Cell Signaling Technology (1 mentions)
3 protocols using cleaved caspase 4
1
Mitochondrial Protein Extraction and Analysis
2
Multimarker Immunofluorescence Staining Protocol
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Primary antibodies used for immunostaining included those for EGFP (Santa Cruz Biotechnology), MMP7 (R & D Systems, Minneapolis, MN, USA), cytokeratin (Leica, Solms, Germany), CD34 (Abcam, Cambridge, MA, USA), GRP78 (Enzo Life Sciences, Lausen, Switzerland), cleaved caspase-3, cleaved caspase-4, cleaved caspase-9, cleaved caspase-12, Cox IV (all from Cell Signaling Technology, Danvers, MA, USA); secondary antibodies included goat anti-mouse HRP (Santa Cruz Biotechnology), goat anti-rabbit HRP (Santa Cruz Biotechnology), donkey anti-goat HRP (Santa Cruz Biotechnology), chicken anti-rat AP (Santa Cruz Biotechnology), chicken anti-rabbit AP (Santa Cruz Biotechnology), chicken anti-mouse AP (Santa Cruz Biotechnology), goat anti-rabbit Alexa 488/594 (Invitrogen), rabbit anti-goat Alexa 594 (Invitrogen), chicken anti-rabbit Alexa 594 (Invitrogen) and chicken anti-mouse Alexa 594 (Invitrogen). Immunostaining was performed using these antibodies. The sections were counter-stained with hematoxylin or nuclear fast red (Vector, Burlingame, CA, USA). For double staining, EGFP, MMP7, cytokeratin, CD34 and PAS staining was performed following TUNEL staining or cleaved caspase-3 staining.
3
Evaluating HePG2 Cell Response to Trinitrotoluene Exposure
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The human hepatoma cell line HePG2 was derived from human hepatocellular carcinoma tissues, which exhibited proliferation kinetics similar to normal liver cells and are widely used to model normal liver cells in vitro. The cell line was kindly provided by the Harbin Medical University.
GRP78, GRP94, cleaved Caspase 4, p-Jun JNK, and CHOP antibodies were obtained from Cell Signaling Technology (Beverly, MA, USA).
HepG2 cells were thawed at 40°C, recovered at 37°C, and grown in culture flasks containing DMEM with 12% FBS at 37°C and 5% CO2. Cells were passaged when they grew to 70–80% confluence. Pure TNT (>99.9% purity) was provided by a machinery factory, and used after recrystallization and purification.
Control were cultured in 10 mL fresh DMEM with 12% FBS. TNT-treatment groups were cultured in 10 mL fresh DMEM with 12% FBS and TNT at 6, 12, and 24 μg/mL for 12, 24, and 48 h.
GRP78, GRP94, cleaved Caspase 4, p-Jun JNK, and CHOP antibodies were obtained from Cell Signaling Technology (Beverly, MA, USA).
HepG2 cells were thawed at 40°C, recovered at 37°C, and grown in culture flasks containing DMEM with 12% FBS at 37°C and 5% CO2. Cells were passaged when they grew to 70–80% confluence. Pure TNT (>99.9% purity) was provided by a machinery factory, and used after recrystallization and purification.
Control were cultured in 10 mL fresh DMEM with 12% FBS. TNT-treatment groups were cultured in 10 mL fresh DMEM with 12% FBS and TNT at 6, 12, and 24 μg/mL for 12, 24, and 48 h.
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