Hrp conjugated goat anti rabbit igg

Manufactured by Wuhan Servicebio Technology

Sourced in China

HRP-conjugated goat anti-rabbit IgG is a secondary antibody used in various immunochemical techniques. It is produced by conjugating horseradish peroxidase (HRP) to goat-derived antibodies that specifically recognize and bind to rabbit immunoglobulin G (IgG) molecules. This conjugated antibody can be used to detect and visualize the presence of rabbit primary antibodies in experimental samples.

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20 protocols using hrp conjugated goat anti rabbit igg

Histological Analysis of Lung Tissue

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After fixed in 4% paraformaldehyde for 7 days, lung tissues were embedded in paraffin, cut into slices. After hematoxylin and eosin (H&E) staining, pathological changes were evaluate and scored on a 0–5 severity scale, according to the thickened alveolar walls, cell aggregation, blocked bronchioles and lung consolidation. Infiltration was also evaluated and scored on a 1–4 severity scale.
For immuno-fluorescence assay, the sections were deparaffinized and rehydrated, followed by Citrate-mediated antigen retrieval. After blocking, the primary antibody Anti-Syndecan-1/CD138 Rabbit pAb (cat. no. GB115052, Servicebio), HRP-conjugated goat-anti-rabbit IgG and CY3-Tyramide (cat. no. G1223, Servicebio) were used, followed by Citrate-mediated antigen retrieval. Then another primary antibody Anti-IL-17 Rabbit pAb (cat. no. GB11110-1, Servicebio), secondary antibody HRP-conjugated goat-anti-rabbit IgG and the corresponding iF488-Tyramide (cat. no. G1231, Servicebio) were used. Cell nuclei were stained using 49, 6-diamino-2-phenylindole (DAPI) (cat. no. G1012, Servicebio). The slide scanner Pannoramic MIDI (3DHISTECH) was utilized to image whole slide.

Li X., Xu M., Yang J., Zhou L., Liu L., Li M., Wang S., Liu M.Q., Huang Z., Zhang Z., Liu S., Hu Y., Lin H., Liu B., Sun Y., Wu Q., Shi Z.L., Lan K., Chen Y., Yan H, & Chen Y.Q. (2024). Nasal vaccination of triple-RBD scaffold protein with flagellin elicits long-term protection against SARS-CoV-2 variants including JN.1. Signal Transduction and Targeted Therapy, 9, 114.

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Immunohistochemical Analysis of Tumor Markers

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The tissues were made into formalin-fixed paraffin-embedded tissue microarrays and slides, after which we performed dewaxing using xylene and hydration using a grade alcohol series. After citric acid solution or EDTA buffer (pH 9.0) were used for antigen retrieval, 3% H₂O₂ was applied for the inhibition of endogenous peroxidase. Then the sections were incubated with 5% BSA for 30 min. Next, HRP-conjugated goat anti-rabbit IgG (1:200, Servicebio) was incubated as the secondary antibody for 50 min. Sections were then stained with 3,3-diaminobenzidine (DAB) (Dako). The primary antibodies used were listed as follows: The anti-biglycan primary antibody (diluted 1:2000, Abcam), anti-CD163 primary antibody (diluted 1:500, Servicebio), anti-FOXP3 primary antibody (diluted 1:500, Abcam). Protein expression was assessed with the value of the mean optical density (MOD) using Image-Pro Plus 6.0 software. The definition of BGN positivity was that the ratio of the value of MOD in the tumor tissue to that in the tumor-adjacent tissue of the same patient was greater than 1. Conversely, if the ratio was less than or equal to 1, it was considered BGN negative.

He Z.X., Zhao S.B., Fang X., E J.F., Fu H.Y., Song Y.H., Wu J.Y., Pan P., Gu L., Xia T., Liu Y.L., Li Z.S., Wang S.L, & Bai Y. (2022). Prognostic and Predictive Value of BGN in Colon Cancer Outcomes and Response to Immunotherapy. Frontiers in Oncology, 11, 761030.

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Evaluation of Oncolytic Adenovirus Therapy

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On day 24, mice from each group were euthanized. The lungs and livers were harvested, and the tumor metastatic lesions in the lung were counted. Then, the lung tissues were processed and stained with H&E. Moreover, the distribution of oncolytic adenoviruses in the lung was analyzed by immunohistochemistry using a mouse anti-adenovirus antibody (Abcam, Cambridge, MA) and a goat anti-human decorin antibody (R&D Systems, Minneapolis, MN). Furthermore, the infiltration of lymphocytes, including CD3⁺ T lymphocytes and macrophages, was also detected by immunohistochemistry using rabbit anti-mouse CD3 (Abcam, Cambridge, MA) and rabbit anti-mouse CD68 (Servicebio, Wuhan, China), respectively. Horseradish peroxidase (HRP)-conjugated rabbit anti-goat immunoglobulin G (IgG) (Servicebio, Wuhan, China) or HRP-conjugated goat anti-rabbit IgG was used as the secondary antibody (Servicebio, Wuhan, China).

Zhang Y., Liu C., Wang T., Kong F., Zhang H., Yi J., Dong X., Duan H., Tao N., Yang Y, & Wang H. (2022). Therapeutic effects of mesenchymal stem cells loaded with oncolytic adenovirus carrying decorin on a breast cancer lung metastatic mouse model. Molecular Therapy Oncolytics, 24, 486-496.

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Histological Analysis of Rat Kidney Apoptosis

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Rat kidneys were dissected and fixed in 4% paraformaldehyde and embedded in paraffin, and 4 μm serial sections were then prepared for histological analysis under a light microscope or fluorescence microscope. TdT-mediated dUTP nick-end labeling (TUNEL) was done with an apoptosis detection kit (Servicebio) following the manufacturer’s instructions. Hematoxylin and eosin (HE) staining and periodic acid-Schiff (PAS)[27 (link)] staining were carried out using standard protocols. A random sample of three glomeruli from each rat was analyzed using image analysis software (Image-Pro plus 6.0) to determine the percentage of PAS-positive areas, expressed as a mesangial index. Immunohistochemical (IHC) analyses were carried out using the rabbit anti-AGEs (1:300; bs-1158R; Bioss, Woburn, MA, United States) primary antibodies. HRP-conjugated goat anti-rabbit IgG (1:500; GB23303; Servicebio) was used to detect primary antibodies. The nuclei were subsequently stained with 3,3’-diaminobenzidine, and three fields of view from each rat were digitized. The integrated optical density from all fields was calculated using Image-Pro Plus 6.0.

Tang L.X., Wei B., Jiang L.Y., Ying Y.Y., Li K., Chen T.X., Huang R.F., Shi M.J, & Xu H. (2022). Intercellular mitochondrial transfer as a means of revitalizing injured glomerular endothelial cells. World Journal of Stem Cells, 14(9), 729-743.

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Inflammasome Activation Pathway Protocol

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AM (P09J8F37712) were obtained from Yuanye Bio-Technology (Shanghai, China), its purity was ≥99.0%; LPS (L4391), MSU (U2875), and phorbol myristate acetate (PMA, P8139) were purchased from Sigma-Aldrich (St. Louis, United States). PI (E24567B117) and hoechst 33,342 (EZ5679A169) were purchased from BioFRoxx (Einhausen, DE). Colchicine (COL, C106740) was from Aladdin (Shanghai, China). TNF-α and IL-1β enzyme-linked immunosorbent assay (ELISA) kits were purchased from Jiancheng Bioengineering Institute (Nanjing, China). LDH and NO assay kits were purchased from Beyotime (Shanghai, China). Antibodies of β-actin, caspase-1, IL-1β, NLRP3, and ASC were obtained from ABclonal (Wuhan, China). FITC-conjugated goat anti-rabbit IgG (GB22303) and HRP-conjugated goat anti-rabbit IgG were bought from Servicebio (Wuhan, China). All cell culture reagents were bought from Hyclone.

Zhang X., Liu Y., Deng G., Huang B., Kai G., chen K, & Li J. (2021). A Purified Biflavonoid Extract From Selaginella moellendorffii Alleviates Gout Arthritis via NLRP3/ASC/Caspase-1 Axis Suppression. Frontiers in Pharmacology, 12, 676297.

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Histological and Immunohistochemical Analysis of Colon Tissue

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In brief, fresh colon tissues were fixed in 4% paraformaldehyde at 4 °C overnight, then subjected to paraffin imbedding sections. For H&E Staining, the sections were stained with haematoxylin and dehydration in graded alcohols and xylene. For immunolabeling, the sections were incubated with indicated primary antibodies: anti-CD3 [1:400, 85061; Cell Signaling]; anti-Ki67 [1:200, GB13030-2; Servicebio], anti-F4/80 [1:200, GB11027; Servicebio], anti-ChgA [1:400, ab45179; Cell Signaling] anti-CAI [1:200, SC39349; Santa Cruz], anti-Lysozyme [1:300, ab108502; Abcam], anti-N2ICD [1:200, YC0069; Immunoway], overnight at 4 °C in the dark. Then, the sections were incubated with either HRP–conjugated Goat anti-Rabbit IgG (1:200, G1215; Servicebio) or HRP–conjugated Goat anti-Mouse IgG (1:200, G1214; Servicebio) for 50 min at 25 °C. The subsequent detection was performed using the standard substrate detection of DAB. TUNEL assay kit was purchased from Abcam (ab66110). Images were taken by using Leica DM6 B Upright Microscope.

Pu Y., Song Y., Zhang M., Long C., Li J., Wang Y., Xu Y., Pan F., Zhao N., Zhang X., Xu Y., Cui J., Wang H., Li Y., Zhao Y., Jin D, & Zhang H. (2021). GOLM1 restricts colitis and colon tumorigenesis by ensuring Notch signaling equilibrium in intestinal homeostasis. Signal Transduction and Targeted Therapy, 6, 148.

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Nrf2-Mediated Antioxidant and Anti-Inflammatory Assay

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Doxorubicin (DOX, #HY-15142), Butylphthalide (NBP, #HY-B0647), ML385 (#HY-100523), Polyethylene glycol 300 (PEG300, #HY-Y0873), and Dimethyl sulfoxide (DMSO, #HY-Y0320) were obtained from MedChemExpress (New Jersey, USA). Anti-Nuclear factor E2-related factor 2 (Nrf2, #GTX103322), anti-heme oxygenase 1 (HO-1, #GTX101147), anti–NF–κB P65 (P65, #GTX102090), and anti-phospho–NF–κB P65 (P–P65, #GTX133899) were purchased from GeneTex (California, USA). Anti-interleukin 6 (IL-6, #YT5348) and antitumor necrosis factor α (TNF-α, #YT4689) were obtained from ImmunoWay (Texas, USA). Anti-IL-1β (#ab283822) was purchased from Abcam (Cambridge, UK). Anti-cleaved caspase3 (C-Caspase3, #9664) was obtained from Cell Signaling Technology (Massachusetts, USA). Anti-NADPH quinone oxidoreductase-1 (NQO1, #A19586), anti-superoxide dismutase-2 (SOD2, #A1340), anti-B cell lymphoma 2 (BCL-2, #A20777), and anti-BCL-2-associated X protein (BAX, #A11931) were purchased from ABclonal Technology (Wuhan, China). Anti-α-Tubulin antibody and HRP conjugated Goat Anti-Rabbit IgG were obtained from Servicebio Technology (Wuhan, China).

Li D., Zhang W., Fu H., Wang X., Tang Y, & Huang C. (2024). DL-3-n-butylphthalide attenuates doxorubicin-induced acute cardiotoxicity via Nrf2/HO-1 signaling pathway. Heliyon, 10(5), e27644.

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Immunohistochemical Analysis of Hippocampal Neuroinflammation

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Brains were rapidly removed, frozen at −80 °C, and cut into 2-mm coronal slices in a rat brain matrix. After 10% formalin fixation, hippocampal tissue samples were processed into paraffin-embedded blocks and 5-μm-thick sections were cut for immunostaining. Hippocampal CA1 region sections were deparaffinized and treated with EDTA antigen repair buffer (pH 8.0). Antibodies used were anti-ionized calcium binding adaptor molecule 1 (IBA-1; anti-Iba1 rabbit pAb, Servicebio, HRP anti-rabbit IgG (HRP-conjugated goat anti-rabbit IgG, Servicebio), anti-glial fibrillary acidic protein (anti-GFAP; anti-GFAP rabbit pAb, Servicebio), and Cy3 anti-rabbit (Cy3-conjugated goat anti-rabbit IgG, Servicebio). Finally, the sections were sealed with the antifluorescence solution (G1401-5ML, Servicebio), and the immunofluorescence images were observed and recorded with fluorescence microscope (NIKON ECLIPSE C1, NIKON DS-U3).

Li P., Huang W., Yan Y.N., Cheng W., Liu S., Huang Y., Chen W., Chen Y.P., Gao Y., Lu W., Xu Y, & Meng X. (2021). Acupuncture Can Play an Antidepressant Role by Regulating the Intestinal Microbes and Neurotransmitters in a Rat Model of Depression. Medical Science Monitor : International Medical Journal of Experimental and Clinical Research, 27, e929027-1-e929027-12.

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Western Blot Analysis of Liver and Cell Proteins

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Liver and cell extracts were obtained using RIPA buffer (Beyotime, Guangzhou, China). After extraction, protein concentrations were determined by BCA. Equal amounts of proteins were separated via 10–12% SDS-PAGE and subsequently electrotransferred to PVDF membranes. After blocking with 5% nonfat milk, the blots were incubated with various primary antibodies, followed by HRP-conjugated secondary antibodies and then developed with ECL reagent (P0018FS, Beyotime, China). The primary antibodies included anti-YSK1 (1:500, Santa Cruz, sc-271196), anti-STRN (6) (1:500, Santa Cruz, sc-136084), anti-AMPK-α1 (1:1000, Beyotime, AF1627), anti-p-AMPK (1:1000, Abcam, ab133448), anti-ATP citrate lyase (1:2000, Abcam, ab40793), and anti-ACC1 (1:1000, Proteintech, 21923-1-AP), anti-beta actin (1:1000, Servicebio, GB11001), anti-E-cadherin (1:5000, proteintech, 20874-1-AP), anti-N-cadherin (1:5000, proteintech, 22018-1-AP), anti-snail (1:500, Proteintech, 13099-1-AP), anti-vimentin (1:2000, Proteintech, 10366-1-AP), HRP-conjugated goat anti-rabbit IgG (1:5000, Servicebio, GB23303), HRP-conjugated goat anti-mouce IgG (1:5000, Servicebio, GB23301). The densities of the proteins were quantified with ImageJ software.

Zhang Y., Xu J., Qiu Z., Guan Y., Zhang X., Zhang X., Chai D., Chen C., Hu Q, & Wang W. (2022). STK25 enhances hepatocellular carcinoma progression through the STRN/AMPK/ACC1 pathway. Cancer Cell International, 22, 4.

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Hippocampal Protein Extraction and Analysis

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Proteins from the hippocampus were extracted by RIPA lysate (Beyotime, Shanghai, China). The primary antibodies used were beta Actin (Servicebio, Wuhan, China), GAD1 and GAD2 (Abcam, Cambridge, MA, USA). Secondary antibody was HRP conjugated Goat Anti-Rabbit IgG (Servicebio, Wuhan, China). Chemiluminescence was performed by Immobilon Western HRP substrate (Millipore, Massachusetts, USA).

Li C., Lin Y., Lin R., Chen Z., Zhou Q., Luo C, & Mo Z. (2023). Host miR-129-5p reverses effects of ginsenoside Rg1 on morphine reward possibly mediated by changes in B. vulgatus and serotonin metabolism in hippocampus. Gut Microbes, 15(2), 2254946.

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