Anti cmaf

Manufactured by Santa Cruz Biotechnology

Sourced in Germany

Anti-CMAF is a laboratory reagent used in research applications. It functions as an antibody that specifically binds to the CMAF protein, which is a transcription factor involved in various cellular processes. This product can be utilized in techniques such as immunoprecipitation, Western blotting, and immunohistochemistry to study the CMAF protein and its interactions.

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3 protocols using anti cmaf

Peripheral Blood T and M Cell Phenotyping

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For surface staining, peripheral blood samples were incubated with anti-CD4, anti-CD4, or F4/80 (eBioscience, San Diego, CA, USA) at 4 °C for 30 min. Then, cells were permeabilized for intracellular staining, which stained with anti-FOXP3, anti-IL-17A, anti-IFNγ, anti-IL-4, anti-inducible nitric oxide synthase (iNOS), and anti-CMAF (Santa Cruz) for Treg, Th17, Th1, Th2, M1 and M2 detections, respectively. Cells were acquired on Canto II flow cytometer (BD Biosciences), and data were analyzed with FlowJo v.9.5.2 software (Tree Star). Lastly, labeled cells were enumerated by Canto II flow cytometer (BD Biosciences, San Jose, CA, USA). Data were analyzed by BD FACSDiva™ Software 6.0 (BD).

Li L.L., Dai B., Sun Y.H, & Zhang T.T. (2020). The activation of IL-17 signaling pathway promotes pyroptosis in pneumonia-induced sepsis. Annals of Translational Medicine, 8(11), 674.

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Chromatin Immunoprecipitation of Transcription Factors

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All antibodies used in these experiments were either conjugated in-house or purchased as indicated. Anti-IL-4 (11B11), anti-IL-12 (C17.18), anti-IFNγ (AN17.18.24), and anti-CD4 (GK1.5) antibodies were purified from hybridoma supernatants at the German Rheumatism Research Center and used at 10 μg/ml final concentration. FITC-conjugated anti-IFNγ (AN18.17.24; BD Pharmingen, Heidelberg, Germany), phycoerythrin-conjugated anti-IL-4 (11B11; BD Pharmingen, and BVD4–1D11; Miltenyi Biotec, Bergisch Gladbach, Germany) were used for intracellular cytokine staining. For the chromatin immunoprecipitation, the following antibodies were used: anti-c-MAF, anti-RNA polymerase II, anti-p300 and anti-STAT6 (polyclonal rabbit IgG; Santa Cruz Biotechnology, Heidelberg, Germany), anti-NFATc2 and -NFATc1 (polyclonal rabbit IgG; ImmunoGlobe Antikörpertechnik GmbH, Himmelstadt, Germany), anti-GATA-3 (mouse monoclonal; Santa Cruz Biotechnology), anti-NF-kB (polyclonal goat IgG; Santa Cruz Biotechnology), anti-Brg1 (rabbit antiserum; Merck-Millipore, Darmstadt, Germany). For the image cytometry, anti-NFATc2 (rabbit monoclonal, clone D4B1, Cell Signaling Technology, Leiden, The Netherlands) and donkey anti-rabbit IgG (coupled to Alexa Fluor 647, Molecular probes A31573, Darmstadt, Germany) were used.

Köck J., Kreher S., Lehmann K., Riedel R., Bardua M., Lischke T., Jargosch M., Haftmann C., Bendfeldt H., Hatam F., Mashreghi M.F., Baumgrass R., Radbruch A, & Chang H.D. (2014). Nuclear Factor of Activated T Cells Regulates the Expression of Interleukin-4 in Th2 Cells in an All-or-none Fashion. The Journal of Biological Chemistry, 289(39), 26752-26761.

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ChIP-seq Analysis of Th17 Cell Transcription Factors

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Cells were polarized in Tr1 conditions for 3 days, fixed with 1% formaldehyde for 10 mins and quenched with 0.125M glycine. Chromatin fraction preparation and chromatin IP was performed using SimpleChIP Enzymatic Chromatin IP (Cell Signaling Technology) according to the manufacturer’s instructions. The following antibodies were used: anti-IRF1 (Santa Cruz sc-640X), anti-BATF (Cell Signaling 8638S), anti-c-Maf (Santa Cruz sc-7866), anti-AhR (Enzo Life Sciences BML-SA210-0025), anti-trimethyl-Histone H3 (Lys4) (Millipore 07-473), anti-trimethyl-Histone H3 (Lys27) (Millipore 07-449), anti-Histone H3 (acetyl K9 (Abcam ab4441). For Re-ChIP, chromatin fraction was prepared as described above, and the chromatin IP was prepared using Re-ChIP-IT kit (Active Motif) according to manufacturer’s instructions. ChIP and Re-ChIP quantitative PCR (qPCR) was run on Viia7 Real-Time PCR System (Applied Biosystems) and relative expression was calculated using Ct values of the samples and inputs. Primers spanning Il17a and IL23r sites were described before^{29 (link)}. The following primer pairs were used for the rest of the targets:

Karwacz K., Miraldi E.R., Pokrovskii M., Madi A., Yosef N., Wortman I., Chen X., Watters A., Carriero N., Awashti A., Regev A., Bonneau R., Littman D, & Kuchroo V.K. (2017). Critical role of the transcription factors IRF1 and BATF in preparing the chromatin landscape during Type 1 regulatory cell differentiation. Nature immunology, 18(4), 412-421.

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