Goat anti rabbit hrp conjugate

Manufactured by Cell Signaling Technology

Sourced in United States

The Goat anti-rabbit HRP conjugate is a secondary antibody that is conjugated to the enzyme horseradish peroxidase (HRP). It is designed to recognize and bind to primary rabbit antibodies, allowing for the detection and visualization of target proteins or antigens in various immunoassays and immunohistochemical applications.

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Lab products found in correlation

Bovine serum albumin (bsa), by Sangon (5 mentions) Anti mettl3, by Abcam (2 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Anti flot1, by Abcam (1 mentions) Anti ki67, by Cell Signaling Technology (1 mentions) Donkey anti mouse alexa fluor 555, by Thermo Fisher Scientific (1 mentions) Rabbit anti vimentin, by Abcam (1 mentions) Chicken anti vimentin, by BioLegend (1 mentions) Mouse anti gfap, by Merck Group (1 mentions) Rabbit anti tomm20, by Abcam (1 mentions) Alexa fluor 555 donkey anti rabbit, by Thermo Fisher Scientific (1 mentions) Horse anti mouse hrp conjugate, by Cell Signaling Technology (1 mentions) Alexa fluor 647 donkey anti rabbit, by Thermo Fisher Scientific (1 mentions) Alexa fluor 488 goat anti chicken, by Thermo Fisher Scientific (1 mentions) Mouse anti ki67, by BD (1 mentions) Anti smad3, by Cell Signaling Technology (1 mentions) Anti yap1, by Cell Signaling Technology (1 mentions) Hrp conjugated anti mouse igg, by Cell Signaling Technology (1 mentions) Anti mettl14, by Merck Group (1 mentions) Anti pcna, by Santa Cruz Biotechnology (1 mentions)

Close spelling variants of this product

Goat anti-rabbit HRP (33 citations) Anti-HRP rabbit (4 citations)

8 protocols using goat anti rabbit hrp conjugate

Quantitative Analysis of Ki-67 and FLOT1 in Bladder Cancer

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Tissue sections of xenograft tumors in nude mice and the same bladder cancer TMAs used in CISH staining were analyzed in this study. TMA was obtained from Xinchao Biotech, Shanghai, China. All the paraffin tissue sections were dewaxed and rehydrated. Antigen retrieval was performed by heating the slides in sodium citrate buffer (10 mM, pH 6.0). After blocking with bovine serum albumin (Sango Biotech, Shanghai, China), the slides were incubated with anti-Ki-67 (Cell Signaling Technology, Beverly, MA, USA), or anti-FLOT1 (Epitomics, Burlingame, CA, USA) overnight at 4 °C. The slides were then incubated with a secondary antibody of goat anti-rabbit HRP conjugate (Cell Signaling Technology, Beverly, MA, USA) for 1 h at room temperature. A DAB solution was used for brown color development. The strength of positivity was semi-quantified by considering both the intensity and proportion of positive cells.

Liang Z., Wang X., Xu X., Xie B., Ji A., Meng S., Li S., Zhu Y., Wu J., Hu Z., Lin Y., Zheng X., Xie L, & Liu B. (2017). MicroRNA-608 inhibits proliferation of bladder cancer via AKT/FOXO3a signaling pathway. Molecular Cancer, 16, 96.

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Antibody Validation for Stem Cell Markers

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Rabbit anti-nestin (for immunofluorescence 1:2500, for western blot 1:2000; BioLegend (San Diego, CA, USA, 839801), mouse anti-GFAP (for immunofluorescence 1:100; Merck (Darmstadt, Germany), MAB360; for western blot 1:250; Dako (Glostrup, Denmark), M0761), chicken anti-vimentin (for immunofluorescence 1:1000; used throughout the study; for western blot 1:2000; BioLegend, 919101), rabbit anti-vimentin (1:200; Abcam (Cambridge, UK), ab45939; used for the comparison in Figure 1), rabbit anti-TOMM20 (1:200; Abcam, ab186734), mouse anti-Ki67 (1:50, BD Biosciences (Franklin Lakes, NJ, USA, 550609), goat anti-chicken Alexa Fluor 488 (1:1000; Thermo Fisher Scientific, (Waltham, MA, USA, A11039), donkey anti-mouse Alexa Fluor 555 (1:1000; Thermo Fisher Scientific, A31570), donkey anti-rabbit Alexa Fluor 647 (1:1000; Thermo Fisher Scientific, A31573), donkey anti-rabbit Alexa Fluor 555 (1:1000; Thermo Fisher Scientific, A31572), rabbit anti-GAPDH–HRP conjugate (1:500; Cell Signaling Technology, (Beverly, MA, USA, 3683), goat anti-rabbit-HRP conjugate (1:1000; Cell Signaling Technology, 7074), and horse anti-mouse-HRP conjugate (1:1000; Cell Signaling, 7076) were used. The specificity of the GFAP, vimentin, and nestin antibodies was previously validated, on tissues/cell cultures from mice carrying null mutations in the respective genes serving as negative controls.

de Pablo Y., Marasek P., Pozo-Rodrigálvarez A., Wilhelmsson U., Inagaki M., Pekna M, & Pekny M. (2019). Vimentin Phosphorylation Is Required for Normal Cell Division of Immature Astrocytes. Cells, 8(9), 1016.

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Immunohistochemical Analysis of SMAD3 Expression

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TMAs were purchased from Xinchao Biotech, Shanghai, China, which contained 46 cases with paired tumor and non-tumor tissues and 13 cases without corresponding non-tumor tissues. All paraffin tissue sections were dewaxed and rehydrated. Antigen retrieval was performed by heating the slides in sodium citrate buffer (10 mM, pH 6.0). After blocking with bovine serum albumin (Sango Biotech, Shanghai, China), the slides were incubated with anti-SMAD3 (Cell Signaling Technology) overnight at 4 °C. The slides were then incubated with a secondary antibody of goat anti-rabbit HRP conjugate (Cell Signaling Technology) for 1 h at room temperature. A DAB solution was used for brown color development. Both the intensity and proportion of positive cells were considered for the semi-quantification of the strength of positivity.

Li J., Xu X., Meng S., Liang Z., Wang X., Xu M., Wang S., Li S., Zhu Y., Xie B., Lin Y., Zheng X., Liu B, & Xie L. (2017). MET/SMAD3/SNAIL circuit mediated by miR-323a-3p is involved in regulating epithelial–mesenchymal transition progression in bladder cancer. Cell Death & Disease, 8(8), e3010-.

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Protein Extraction and Western Blot Analysis

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Protein was extracted from cells or tissues using T-per tissue protein extraction reagent (catalog #78510) lysis buffer. Concentration was measured using the Bradford Assay reagent. 10-40 ug of protein was loaded on SDS-polyacrylamide gel and transferred to a nitrocellulose membrane. The membrane was blotted with anti-Mettl3 (#ab195352 Abcam, #MA5-27527 Invitrogen), anti-Mettl14 (#HPA038002, Sigma), anti-GFP (#GFP-1020 Aves Lab), anti-HA (#3724, Cell Signaling), anti-c-Myc (#ab185656 Abcam, #OP10-200ug Sigma), anti-Avpr2 (#AB1797P, EMD Millipore), anti-Yap1 (#4912, Cell Signaling), anti-pCREB (#9198S, pCreb1), anti-PCNA (#sc-7907, Santa Cruz), or anti-puromycin (#PMY-2A4, Developmental Studies Hybridoma Bank) antibodies. All the primary antibodies were used at 1:1000 dilution, except for anti-puromycin, which was used at 1:50 dilution. Goat anti-rabbit-HRP conjugate (#7074S, Cell Signaling) or anti-mouse HRP-conjugated IgG (#7076S, Cell Signaling) was used as the secondary antibody. HRP-conjugated Actin antibody (#A3865, Sigma) was used at 1:40,000 dilution for measuring total protein. The blots were developed using the X-ray film developer or the Bio-rad digital imager. The protein bands were quantified using the Imagelab software from Bio-rad. Each western blot was repeated at least three times.

Ramalingam H., Kashyap S., Cobo-Stark P., Flaten A., Chang C.M., Hajarnis S., Hein K.Z., Lika J., Warner G.M., Espindola-Netto J.M., Kumar A., Kanchwala M., Xing C., Chini E.N, & Patel V. (2021). A methionine-Mettl3-N6-methyladenosine axis promotes polycystic kidney disease. Cell metabolism, 33(6), 1234-1247.e7.

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Immunohistochemical Analysis of Cancer Markers

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The paraffin tissue sections were subjected to dewaxing and rehydration procedures. After performing antigen retrieval, the slides were subjected to blocking using bovine serum albumin (Sango Biotech, Shanghai, China). Subsequently, the slides were incubated overnight at 4 °C with c-Fos Monoclonal antibody (Proteintech, 66590-1-Ig), Vimentin Polyclonal antibody (Proteintech, 10366-1-AP), Anti-E-cadherin antibody (Abcam, ab231303), PCNA Polyclonal antibody (Proteintech, 10205-2-AP), and HIF-1 alpha Polyclonal antibody (Proteintech, 20960-1-AP). Subsequently, the samples were incubated with a secondary antibody of goat anti-rabbit HRP conjugate (Cell Signaling Technology, Beverly, MA, USA) at 25 °C for 1 h. A DAB solution was employed to facilitate brown color development. The regions of interests (ROIs) were analyzed using IHC Profiler [20 (link)].

Fei X., Liu J., Xu J., Jing H., Cai Z., Yan J., Wu Z., Li H., Wang Z, & Shen Y. (2024). Integrating spatial transcriptomics and single-cell RNA-sequencing reveals the alterations in epithelial cells during nodular formation in benign prostatic hyperplasia. Journal of Translational Medicine, 22, 380.

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Immunohistochemical Evaluation of YTHDF2

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All paraffin tissue sections obtained from above TMAs were dewaxed and rehydrated. Antigen retrieval was performed by heating the slides in sodium citrate buffer (10 mM, pH 6.0). After blocking with bovine serum albumin (Sango Biotech, Shanghai, China), the slides were incubated with anti-YTHDF2 (Proteintech, Chicago, IL, USA) overnight at 4 °C. The slides were then incubated with a secondary antibody of goat antirabbit HRP conjugate (Cell Signaling Technology, Beverly, MA, USA) for 1 h at room temperature. A DAB solution was used for brown color development. HScores system was used to semi-quantify the strength of positivity which considered the intensity of the staining and the percentage of positive cells per the formula: HScores = 1 × (% light staining) + 2 × (% moderate staining) + 3 × (% strong staining), and HScores values range from 0 to 300.

Li J., Meng S., Xu M., Wang S., He L., Xu X., Wang X, & Xie L. (2017). Downregulation of N6-methyladenosine binding YTHDF2 protein mediated by miR-493-3p suppresses prostate cancer by elevating N6-methyladenosine levels. Oncotarget, 9(3), 3752-3764.

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GATA3 and AKT Signaling Pathway Profiling

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Whole cell lysates were prepared using RIPA buffer with quantification using the Pierce™ BCA Protein assay kit (ThermoFisher). Lysates were resolved by SDS-PAGE using 10% acrylamide gels and proteins were transferred onto polyvinylidene fluoride (PVDF) membrane by semi-dry blotting. The following antibodies were used in this study: goat polyclonal CTSV (BioTechne, AF1080), goat polyclonal GAPDH (BioTechne, AF5718), mouse monoclonal GATA3 (BioTechne, MAB6330), rabbit monoclonal pSer308 GATA3 (Abcam, ab186371), mouse monoclonal pThr308 Akt (BioTechne, MAB7419), rabbit polyclonal pSer473 Akt (Cell Signaling, 9271), mouse monoclonal Akt (BD Transduction Laboratories, P78020), rabbit polyclonal pSer9 GSK-3β (BioTechne, AF1590), mouse monoclonal GSK-3β (BioTechne, MAB25063) and mouse monoclonal FLAG M2 (Sigma-Aldrich, F3165). Secondary antibodies used were donkey anti-goat HRP conjugate (BioTechne, HAF109), goat anti-mouse HRP conjugate (BioRad, 172-1011) and goat anti-rabbit HRP conjugate (Cell Signaling, 70745). Protein visualisation was undertaken using Luminata™ Forte chemiluminescent detection reagent (Merck Millipore) and imaged using a Molecular Imager ChemiDoc XRS+ Imaging System (BioRad).

Sereesongsaeng N., McDowell S.H., Burrows J.F., Scott C.J, & Burden R.E. (2020). Cathepsin V suppresses GATA3 protein expression in luminal A breast cancer. Breast Cancer Research : BCR, 22, 139.

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Immunohistochemical Analysis of METTL3 in Prostate Cancer

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The PCa TMA was purchased from Xinchao Biotech, Shanghai, China. It contained three normal prostate tissues, eight adjacent tissues of PCa, and forty-nine PCa tissues. All para n tissue sections obtained from the TMA were dewaxed and rehydrated. After Antigen retrieval, blocking with bovine serum albumin (Sango Biotech, Shanghai, China), the slides were incubated with anti-METTL3 (Abcam, Cambridge, Massachusetts, US) overnight at 4 °C. Then, they were incubated with a secondary antibody of goat antirabbit HRP conjugate (Cell Signaling Technology, Beverly, MA, USA) for 1 h at 25℃. A DAB solution was used for brown color development.

Li H., Fei X., Shen Y., & Wu Z. (2022). Prognostic Value and Possible Mechanism of m6A Methyltransferase METTL3 in Prostate Cancer.

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