Anti ifn β

Manufactured by Abcam

Sourced in United Kingdom

Anti-IFN-β is a primary antibody that recognizes and binds to interferon-beta (IFN-β), a cytokine involved in immune responses. This antibody can be used for the detection and analysis of IFN-β in various applications, such as Western blotting, ELISA, and immunohistochemistry.

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7 protocols using anti ifn β

Immunoblot Analysis of Innate Immunity Proteins

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For immunoblotting analysis, lung samples and cell lysates lysed by RIPA buffer (Beyotime, China) were resolved by SDS–PAGE and transferred to the polyvinylidene fluoride membrane (Millipore, United States). Immunoblots were visualized by the ECL system (Fdbio Science, China). The following antibodies were used in immunoblotting analysis: antibodies for anti-IFITM3 (1:2,000), anti-MFN2 (1:2,000), anti-IFN-β (1:500) antibody and anti-IL-1β (1:1,000) were from Abcam. β-actin antibody (1:2,000) was purchased from Fude Biotechnology. Anti-phospho-IRF3 (1:1,000), and anti-TNF-α (1:2,000) antibodies were purchased from Cell Signaling Technology. Anti-MAVS (1:500) antibodies were obtained from Proteintech Group. β-actin was used as an internal control and the relative densities of the measured protein were quantified by image J software.

Luo Z., Liu L.F., Wang X.H., Li W., Jie C., Chen H., Wei F.Q., Lu D.H., Yan C.Y., Liu B., Kurihara H., Li Y.F, & He R.R. (2019). Epigoitrin, an Alkaloid From Isatis indigotica, Reduces H1N1 Infection in Stress-Induced Susceptible Model in vivo and in vitro. Frontiers in Pharmacology, 10, 78.

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Western Blot Immunodetection Protocol

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The cells or tissues were lysed, and the total protein concentration was measured using the BCA assay. Equal quantities of proteins were resolved using SDS-PAGE and then transferred onto a PVDF membrane (Merck Millipore, Billerica, MA, USA). Next, the membrane was blocked at 37 °C for 2 h and then incubated with the primary antibody (anti-COLII, anti-ACAN, anti-NF-κB, anti-IFN-β, anti-GAPDH [all from Abcam, Cambridge, MA, USA], or anti-Trim14 [Novus Biologicals] antibody) at 4 °C overnight. The membrane was washed three times with PBST (PBS with Tween 20, Solarbio) and then incubated with the horseradish-peroxidase-labeled anti-mouse or anti-rabbit IgG secondary antibody (Jackson Immunoresearch, West Grove, PA, USA) (diluted 1: 10,000 with PBST) at 37 °C for 2 h. Finally, the membrane was washed five times with PBST and developed using chemiluminescence (BeyoECL Star [Beyotime, Shanghai, China]). The results were analyzed using the ImageJ software (NIH, Bethesda, MA, USA).

Wang H., Shu J., Zhang C., Wang Y., Shi R., Yang F, & Tang X. (2022). Extracellular Vesicle-Mediated miR-150-3p Delivery in Joint Homeostasis: A Potential Treatment for Osteoarthritis?. Cells, 11(17), 2766.

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Antibody Validation for Western Blot and Immunofluorescence

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Anti-NF-κB, anti-IRF3, anti-TBK1, anti-IFN-β, anti-GAPDH, anti-pNF-κB, anti-pTBk1, anti-pIRF3 and anti-DDDDK for WB and immunofluorescence (IF) assays were purchased from Abcam (Abcam, Cambridge, UK). Anti-HA and anti-Myc for WB and immunofluorescence (IF) assays were purchased from Thermo Fisher (ThermoFisher, Waltham, MA, USA). Anti-J-2 for IF assays were purchased from Scicons (Scicons, Szirák, Hungary). Secondary antibodies labeled with Alexa Fluor 488, Alexa Fluor 555, Alexa Fluor 594 or Alexa Fluor 647 for the IF assay were purchased from Abcam (Abcam, Cambridge, UK). Goat anti-mouse IgG H&L (HRP) and goat anti-rabbit IgG H&L were purchased from Abcam (Abcam, Cambridge, UK) and used in the WB assays.

Yang Z., Li J., Li J., Zheng H., Li H., Lai Q., Chen Y., Qin L., Zuo Y., Guo L., Shi H, & Liu L. (2022). Engagement of the G3BP2-TRIM25 Interaction by Nucleocapsid Protein Suppresses the Type I Interferon Response in SARS-CoV-2-Infected Cells. Vaccines, 10(12), 2042.

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Protein Extraction and Western Blot Analysis

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The cells were lysed in RIPA buffer (Servicebio, China) with a protease inhibitor (PMSF, Servicebio, China) using routine procedures to extract total proteins. Lysates were centrifuged at 12,000 rpm for 10 min, followed by the transfer of supernatants to fresh EP tubes. The BCA protein assay was used for protein quantification. Each sample (25 μg) was loaded onto a 12.5 % SDS-PAGE. The membrane was treated with 5 % non-fat milk for 1 h and then washed three times with TBST (Servicebio, China). After washing, membranes were left overnight at 4 °C with anti–IFN–β (1:1000, Abcam, UK) and anti–HIF–1 alpha (1:1000, Abcam, UK) antibodies. After washing, the membranes were incubated for 2 h with anti-rabbit secondary antibodies. Membranes were developed using enhanced chemiluminescence (ECL).

Xiong S., Zhang Y., Zeng J., Zhou J., Liu S., Wei P., Liu H., Yi F., Wan Z., Xiong L., Zhang B, & Li J. (2023). DLP fabrication of HA scaffold with customized porous structures to regulate immune microenvironment and macrophage polarization for enhancing bone regeneration. Materials Today Bio, 24, 100929.

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Protein Expression Analysis of Immune Signaling

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Cells were homogenized in radio-immunoprecipitation assay buffer supplemented with Pierce™ EDTA-free Protease Inhibitor Tablets and Pierce™ Phosphatase Inhibitor Mini Tablets (all Thermo Fisher Scientific) for 30 min at 4 °C, and then centrifuged at 10,000 g for 15 min at 4 °C. The protein concentrations were determined by a bicinchoninic acid Protein Concentration Determination Kit (Thermo Fisher Scientific). The proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein expression was determined using anti-p-STAT1, anti-STAT1, anti-TBK1, anti-IRF3, anti-p-TBK1, anti-p-IRF3, anti-GAPDH (all Cell Signaling Technology), anti-IFN-β, or anti-GBP2 (all Abcam, Cambridge, UK) antibodies at 4 °C overnight. Finally, membranes were incubated with appropriate secondary antibodies (Abcam) and scanned by a Bio-Rad ChemiDoc MP imager (Bio-Rad). The antibodies used were listed in Additional file 1: Table S1.

Du C.H., Wu Y.D., Yang K., Liao W.N., Ran L., Liu C.N., Zhang S.Z., Yu K., Chen J., Quan Y., Chen M., Shen M.Q., Tang H., Chen S.L., Wang S., Zhao J.H., Cheng T.M, & Wang J.P. (2023). Apoptosis-resistant megakaryocytes produce large and hyperreactive platelets in response to radiation injury. Military Medical Research, 10, 66.

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Decitabine Induces ERV-3 and IFNβ

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MOLM-14 and MV4–11 were treated with low-dose decitabine (25nM and 50nM) daily X 3 days. Cells were assessed for ERV-3 (anti-ERV-3 Everest Biotech, Oxfordshire, UK, 30 kD) and IFNβ (anti-IFNβ Abcam, Cambridge, UK, 22 kD) expression.

Nahas M.R., Stroopinsky D., Rosenblatt J., Cole L., Pyzer A.R., Anastasiadou E., Sergeeva A., Ephraim A., Washington A., Orr S., McMasters M., Weinstock M., Jain S., Leaf R.K., Ghiasuddin H., Rahimian M., Liegel J., Molldrem J.J., Slack F, & Avigan D. (2019). Hypomethylating agent alters the immune microenvironment in AML and enhances the immunogenicity of a DC/AML vaccine. British journal of haematology, 185(4), 679-690.

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Multiparameter Flow Cytometry Analysis

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Anti-mouse CD45 (30-F11), Ly-6G (1A8), Ly-6C (AL-21), CD11b (M1/70), CD11c (HL3), F4/80 (BM8), CD49b (DX5), CD3ε (145–2C11), CD4 (GK1.5), CD8α (Ly-2, 53–6.7), and CD19 (1D3) flow cytometry antibodies were purchased from BD Biosciences (San Jose, CA). Flow cytometry analysis was performed 48 hr after treatment. Blood samples were obtained via retro-orbital bleeding and mice were euthanized immediately afterwards and tumors and spleens were harvested. Organs were gently disrupted into single-cell suspensions in progressive steps. Cells were stained to identify immune cell populations with a blocking step using anti-mouse CD18/CD32 and analyzed using a BD FACS LSR II flow cytometer (Becton Dickinson, Franklin Lakes, NJ). FlowJo software was used to analyze data. For intracellular IFNβ staining, surface-stained cells were fixed and permeabilized (Fix/Perm Wash Buffer, BioLegend, San Diego, CA), stained with anti-IFNβ (Abcam, Cambridge, MA), and followed by secondary antibody staining.

Atukorale P.U., Raghunathan S.P., Raguveer V., Moon T.J., Zheng C., Bielecki P.A., Wiese M.L., Goldberg A.L., Covarrubias G., Hoimes C.J, & Karathanasis E. (2019). Nanoparticle encapsulation of synergistic immune agonists enables systemic co-delivery to tumor sites and interferon β-driven anti-tumor immunity. Cancer research, 79(20), 5394-5406.

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