Total protein lysates were obtained from H9c2 cells using the Nuclear and Cytoplasmic Protein Extraction Kit (Thermo Fisher Scientific, Waltham, USA). Approximately 40 μg of proteins was separated on 12% SDS-polyacrylamide gel (SDS-PAGE), and the separated proteins in the gel were transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, MA, USA). The blot was blocked with 5% non-fat dry milk in TBST (Tris-buffered saline/0.1% Tween-20, pH 7.4) for 1 hour at room temperature and incubated overnight with primary antibodies against GPX4 (1:1000) PAI-1 (1:1000), p21 (1:1000), SIRT1 (1:1000) and GAPDH (1:1000) (Abcam, Cambridge, MA). A horseradish peroxidase-conjugated antibody against rabbit IgG (1:5000, Abcam, Cambridge, MA) was used as a secondary antibody. Blots were reacted with the ECL reagents (Amersham Pharmacia Biotech, Inc, USA) and exposed to Tanon 5200-multi chemiluminescent imaging system to detect protein expression. Three independent assays were performed for each group.
Horseradish peroxidase conjugated antibody against rabbit igg
Manufactured by Abcam
Sourced in United States
Horseradish peroxidase-conjugated antibody against rabbit IgG is a secondary antibody that binds to rabbit immunoglobulin G (IgG). The horseradish peroxidase enzyme is conjugated to the antibody, allowing for the detection of target proteins in various immunoassays, such as Western blotting and enzyme-linked immunosorbent assay (ELISA).
Automatically generated - may contain errors
Lab products found in correlation
Gapdh, by Abcam (4 mentions)
Pvdf membrane, by Merck Group (3 mentions)
Nuclear and cytoplasmic protein extraction kit, by Thermo Fisher Scientific (2 mentions)
Nuclear and cytoplasmic protein extraction kit, by Beyotime (2 mentions)
Pai 1, by Abcam (1 mentions)
Sirt1, by Abcam (1 mentions)
Ecl reagent, by Cytiva (1 mentions)
Lc3 1 2, by Abcam (1 mentions)
P nf κb, by Abcam (1 mentions)
Myd88, by Abcam (1 mentions)
Beclin 1, by Abcam (1 mentions)
Caspase 3, by Abcam (1 mentions)
Enhanced chemiluminescence reagent, by Cytiva (1 mentions)
Polyvinylidene difluoride membrane, by Merck Group (1 mentions)
P mtor, by Abcam (1 mentions)
4 protocols using horseradish peroxidase conjugated antibody against rabbit igg
1
Protein Expression Analysis in H9c2 Cells
2
Protein Expression in Lung Tissue
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Inflammatory and autophagy-related proteins were evaluated by Western blot. Proteins were extracted from the lung tissues using the Nuclear and Cytoplasmic Protein Extraction Kit (Beyotime, China). Approximately 40 μg of protein was loaded and separated with the 12% SDS-polyacrylamide gel (SDS-PAGE) and then transferred to polyvinylidene difluoride (PVDF) membrane (Millipore, MIT, USA). The membrane was incubated with 5% nonfat dry milk in TBST (Tris-buffered saline/0.1% Tween-20, pH 7.4) for 1 h at room temperature, followed by incubation overnight with primary rabbit anti-mouse antibodies to NF-κB (1 : 1000, Abcam, USA), p-NF-κB (1 : 1000, Abcam, USA), TLR1 (1 : 1000, Abcam, USA), TLR2 (1 : 1000, Abcam, USA), TLR3 (1 : 1000, Abcam, USA), TLR4 (1 : 1000, Abcam, USA), Myd88 (1 : 1000, Abcam, USA), ATG5 (1 : 1000, Abcam, USA), LC3I/II (1 : 1000, Abcam, USA), Beclin1 (1 : 1000, Abcam, USA), and GAPDH (1 : 1000, Abcam, USA). A horseradish peroxidase-conjugated antibody against rabbit IgG (1 : 5000, Abcam, USA) was used as a secondary antibody. Blots were incubated with the ECL reagents (Beyetime, Jiangsu Province, China) and exposed to Tanon 5200-multi to detect protein expression. Three independent assays were performed.
3
Investigating Protein Expression in Cisplatin-Resistant KYSE-410 Cells
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The KYSE-410-CisRs were incubated for 24 h after being mixed with different drugs, and proteins were isolated from each cell line using the Nuclear and Cytoplasmic Protein Extraction Kit (Beyotime, China). Approximately 40 µg of protein was separated on 12% SDS-polyacrylamide gel (SDS-PAGE), and the gel was placed on a polyvinylidene difluoride (PVDF) membrane (Millipore, MIT, MA, USA). The membrane was blocked with 5% nonfat dry milk in Tris-buffered saline/0.1% Tween-20 (TBST) (pH 7.4) for 1 h at room temperature and followed by overnight incubation with primary rabbit anti-human antibodies to Sirt1 (1:1,000), p53 (1:1,000), p21 (1:1,000), caspase3 (1:1,000), and GAPDH (1:1,000) (Abcam, USA). A horseradish peroxidase-conjugated antibody against rabbit IgG (1:5,000, Abcam, USA) was used as a secondary antibody. Incubation of the blots was completed with ECL reagents (Beyetime, China), while protein expression detection was completed by exposure to Tanon 5200-multi. Three independent assays were performed.
4
Isolation and Western Blot Analysis of Proteins
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Total proteins were isolated from tissues or cells using a Nuclear and Cytoplasmic Protein Extraction Kit (Thermo Fisher Scientific, Waltham, MA, USA). Approximately 40 μg of protein was separated on a 12% SDSpolyacrylamide gel, and the gel was transferred to a polyvinylidene difluoride membrane (Millipore, MIT, USA). The membrane was blocked with 5% nonfat dry milk in Tris-buffered saline/0.1% Tween-20, pH 7.4, for 1 h at room temperature and incubated overnight with primary rabbit anti-human antibodies against TNF-α (1:1000), human leukocyte antigen-DR (HLA-DR) (1:1000), arginase-1 (Arg-1) (1:1000), CD163 (1:1000), CD206 (1:1000), IL-4 (1:1000), AMP-activated protein kinase (AMPK) (1:1000), p-AMPK (1:1000), hypoxia-inducible factor-1α (HIF-1α) (1:1000), protein kinase B (AKT) (1:1000), p-AKT (1:1000), mechanistic target of rapamycin (mTOR) (1:1000), p-mTOR (1:1000), and GAPDH (1:1000) (Abcam, Cambridge, MA, USA). A horseradish peroxidase-conjugated antibody against rabbit IgG (1:5000; Abcam, Cambridge, MA, USA) was used as the secondary antibody. The blots were incubated with enhanced chemiluminescence reagents (Amersham Pharmacia Biotech, Inc., USA) and exposed to a Tanon 5200-multi to detect protein expression. Three independent assays were performed for each sample.
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