Secondary horseradish peroxidase hrp conjugated antibodies

Manufactured by Agilent Technologies

Secondary horseradish peroxidase (HRP)-conjugated antibodies are laboratory reagents used to detect and quantify specific target proteins or antigens in various analytical techniques, such as Western blotting and enzyme-linked immunosorbent assays (ELISAs). These antibodies are conjugated with the enzyme horseradish peroxidase, which can catalyze a colorimetric or chemiluminescent reaction, enabling the visualization and quantification of the target analyte.

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Horseradish peroxidase (94 citations) Horseradish peroxidase (HRP) (20 citations) HRP-conjugated antibodies (20 citations) Secondary horseradish peroxidase-conjugated antibodies (3 citations) Secondary antibodies conjugated to horseradish peroxidase (HRP) (2 citations)

2 protocols using secondary horseradish peroxidase hrp conjugated antibodies

Western Blot Analysis of Acetylated Tubulin

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Cells were treated with or without HSV1716 at 10 PFU/cell and harvested at 24 or 48 hr, and cell lysates were obtained using RIPA buffer containing 25 μL/mL protease inhibitor (Sigma-Aldrich). Proteins (20–40 μg loaded per lane) were separated by SDS-PAGE and transferred to nitrocellulose. Antibodies used were mouse anti-acetylated tubulin (1:5,000 dilution; Sigma-Aldrich), rabbit anti-β-actin (1:2,000 dilution; Abcam), rabbit anti-GSK-3β (1:1,000 dilution; Cell Signaling Technology), rabbit anti-phospho GSK-3β (Ser 9) (1:100 dilution; Cell Signaling Technology), rabbit anti-GSK3 (∝+β) (phospho Y216+Y279; 1:1,000 dilution; Abcam), and secondary horseradish peroxidase (HRP)-conjugated antibodies per the manufacturer’s instructions (Dako and Thermo Fisher Scientific). Proteins were detected by the addition of SuperSignal West Pico Chemiluminescent substrate (Thermo Fisher Scientific) and were visualized using the ChemiDoc MP imaging system (Bio-Rad) and Image Lab (Bio-Rad) software. Protein band intensity was analyzed using ImageJ software. The intensity of each band was normalized to the corresponding β-actin control.

Cockle J.V., Brüning-Richardson A., Scott K.J., Thompson J., Kottke T., Morrison E., Ismail A., Carcaboso A.M., Rose A., Selby P., Conner J., Picton S., Short S., Vile R., Melcher A, & Ilett E. (2017). Oncolytic Herpes Simplex Virus Inhibits Pediatric Brain Tumor Migration and Invasion. Molecular Therapy Oncolytics, 5, 75-86.

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Antibody-based immunoblotting protocol

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Sample preparation and immunoblotting were carried out as previously described^{25 (link)}. The following antibodies from Cell Signalling Technologies (MA, US) were used: anti-phospho-Stat3 (Tyr705: 9131), anti-total-Stat3 (9132), anti-phospho-Akt (Ser473: 9271), anti-total-Akt (9272), anti-phospho-Gsk3β (9331), anti-GSK3β (9315), anti-Akt1 (2967), anti-Akt2 (2964), anti-Akt3 (4059) and anti-Lamin A/C (2032). The following antibodies from Abcam (Cambridge, UK) were used: anti-cathepsin B (ab33538), anti-Lamp2 (ab13524), anti-Tubulin (ab6160) and anti-β-actin (ab8227). The following antibodies from Santa Cruz Biotechnology (CA, US) were used: anti-C/ebpα (sc-61), anti-histone H3 (sc8654), anti-Bax (sc-7480), anti-Bcl-2 (sc7382) and anti-Bcl-xL (sc-634). Other commercial antibodies used were: anti-cathepsin L (MAB9521), anti-cathepsin L (AF1515, used for the immunodetection of cathepsin L in the cathepsin L knockout and control glands) and anti-Bid (MB860) from R&D Systems (MN, US), anti-pan-p85 (Millipore, 06-496, also detects p50α/p55α subunits), anti-cytochrome c (65981A) and anti-E-cadherin (610182) from BD biosciences. All antibodies were used at a standard dilution of 1:1,000. Secondary horseradish peroxidase (HRP)-conjugated antibodies were purchased from Dako (Glostrup, Denmark).

Pensa S., Neoh K., Resemann H.K., Kreuzaler P.A., Abell K., Clarke N.J., Reinheckel T., Kahn C.R, & Watson C.J. (2014). The PI3K regulatory subunits p55α and p50α regulate cell death in vivo. Cell Death and Differentiation, 21(9), 1442-1450.

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