Ab83053

Manufactured by Abcam

Ab83053 is a primary antibody that recognizes the Histone H3 (tri methyl K27) protein. It is designed for use in applications such as immunohistochemistry and western blotting.

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5 protocols using ab83053

Immunofluorescence Analysis of Muscle Tissue

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IF was performed on frozen muscle cross sections that were postfixed after sectioning in precooled acetone (−20°C) for 10 min and placed at room temperature for 30 min. Sections were washed twice in PBS and then blocked by incubating the sections with blocking buffer for 1 h at room temperature. The sections were washed with PBS and incubated with the primary antibodies against dystrophin (1:100; MANDRA11; DSHB), IL6R (1:500; ab83053; Abcam), and CD31 (1:500; ab24590; Abcam) diluted in blocking buffer overnight at 4°C in a humid chamber. Sections were washed, and the secondary antibody was applied for dystrophin (AlexaFluor 594 florescent secondary antibody; A-11032; Thermo Fisher), IL6R (AlexaFluor 594 florescent secondary antibody; A21209; Thermo Fisher), and CD31 (AlexaFluor 488 florescent secondary antibody; A11001; Thermo Fisher) at a dilution of 1:1,000 in blocking buffer for 1 h at room temperature. Sections were washed and incubated with DAPI (268298; EMD Millipore) for 1 min to visualize nuclei. Sections were washed, and coverslips were applied using Prolong Gold Antifade mounting media.

Rupert J.E., Narasimhan A., Jengelley D.H., Jiang Y., Liu J., Au E., Silverman L.M., Sandusky G., Bonetto A., Cao S., Lu X., O’Connell T.M., Liu Y., Koniaris L.G, & Zimmers T.A. (2021). Tumor-derived IL-6 and trans-signaling among tumor, fat, and muscle mediate pancreatic cancer cachexia. The Journal of Experimental Medicine, 218(6), e20190450.

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Regulation of Cardiac Fibrosis by IL-6

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IL-6Rα expression (ab83053, Abcam) was evaluated in discoidin domain receptor 2+ (DDR2; ab76967, Abcam) CFs using immunohistochemistry. The influence of IL-6 on murine specific matrix metalloproteinase 9 (MMP-9) was quantified using media conditioned by murine CFs after 48 h of 1% oxygen and 1% serum direct co-culture with SCR or shIL-6-transduced human EDCs (MMPT90, R&D Systems). The effect of IL-6 on CFs and cardiac macrophages in vivo was assessed 28 days post-infarct using histological analysis of myocardial sections for alpha smooth muscle actin (αSMA; ab66133, Abcam), the myofibroblast marker CD68 (ab955, Abcam) or M2 macrophage marker CD206 (ab64693, Abcam).

Mayfield A.E., Kanda P., Nantsios A., Parent S., Mount S., Dixit S., Ye B., Seymour R., Stewart D.J, & Davis D.R. (2017). Interleukin-6 Mediates Post-Infarct Repair by Cardiac Explant-Derived Stem Cells. Theranostics, 7(19), 4850-4861.

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Protein Expression Analysis by Western Blot

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The protein concentration was determined by bicinchoninic acid protein assay. Equal amount of protein was used, and Western blotting was performed as previously described (66 (link)). Primary antibodies included anti-CD9 antibody (ab92726, Abcam, MA), anti-CD63 antibody (216130, Abcam, MA), anti-CD81 antibody [10037, Cell Signaling Technology (CST)], anti–annexin A1 antibody (3299, CST), anti-HSP90 antibody (4877, CST), anti–IL-6R antibody (ab83053, Abcam, MA), anti-Stat3 antibody and anti-p-Stat3 antibody (9139S and 9145S, CST), anti–IL-6 antibody (12912, CST), anti–nuclear factor κB antibody (8242S, CST), anti–TNF-α antibody (ab1793, Abcam, MA), anti–IL-1b antibody (NB600-633, Novus), anti-CD86 antibody (ab119857, Abcam, MA), anti-Arg1 antibody (sc-18355, Santa Cruz Biotechnology, CA), and anti–β-actin antibody (ab28696, Abcam, MA). The gray intensity of protein blots was measured using ImageJ software (NIH).

Zhou T., He C., Lai P., Yang Z., Liu Y., Xu H., Lin X., Ni B., Ju R., Yi W., Liang L., Pei D., Egwuagu C.E, & Liu X. (2022). miR-204–containing exosomes ameliorate GVHD-associated dry eye disease. Science Advances, 8(2), eabj9617.

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Signaling Pathway Protein Analysis

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Antibodies to IL-6 (AB6672) and IL-6R (AB83053), STAT3-unphosphorylated (AB68153), STAT3 Serine 727 phosphorylation (AB86430), STAT3 Tyrosine 705 phosphorylation (AB76315), JAK2-unphosphorylated (AB98031 and AB108596), JAK2 phosphorylated-Y1007+Y1008 (AB68268), NF-κB-p105/p50 (AB32360), AKT1 (AB32505), and GAPDH (AB9485) were obtained from Abcam (Cambridge, MA). All HRP-conjugated secondary antibodies were from Invitrogen.

Mishra V., DiAngelo S.L, & Silveyra P. (2016). Sex-specific IL-6-associated signaling activation in ozone-induced lung inflammation. Biology of Sex Differences, 7, 16.

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Comprehensive Protein Expression Analysis

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ACTA2 (ab7817, Abcam, 1:1000 for western blot and 1:500 for operetta assay), Albumin (ab207327, Abcam, 1:100 for IF and flow cytometry), Cleaved Caspase-3 (9664, CST, 1:1000), Caspase-3 (9662, CST, 1:1000), Collagen I (ab34710, Abcam, 1:500), phospho-ERK1/2 (4370, CST, 1:1000), ERK1/2 (4695, CST, 1:1000), GAPDH (2118, CST, 1:1000), gp130 (human, PA5-28932, Thermo Fisher, 1:100), gp130 (mouse, PA5-99526, Thermo Fisher, 1:100), gp130 (extracellular, PA5-77476, Thermo Fisher, 1:1000), IgG (11E10, Aldevron), IL6 (AF506, R&D Systems, 1:1000), IL6R (flow cytometry, ab222101, Abcam, 1:100), IL6R (human, IHC and IF, MA1-80456, Thermo Fisher, 1:100), IL6R (mouse, ab83053, Abcam, 1:100), IL11 (X203, Aldevron), IL11RA (inhibition study, X209, Aldevron), IL11RA (IHC, IF, flow cytometry, ab125015, Abcam, 1:100), IL11RA (western blot, sc-130920, Santa Cruz, 1:1000), phospho-JNK (4668, CST, 1:1000), JNK (9252, CST, 1:1000), NOX4 (MA5-32090, Invitrogen, 1:1000), phospho-STAT3 (4113, CST, 1:1000), STAT3 (4904, CST, 1:1000), mouse Alexa Fluor 488 secondary antibody (ab150113, Abcam, 1:200), mouse HRP (7076, CST, 1:2000), rabbit Alexa Fluor 488 secondary antibody (ab150077, Abcam, 1:200), rabbit HRP (7074, CST, 1:2000), rat Alexa Fluor 488 secondary antibody (ab150157, Abcam, 1:200), rat HRP (31470, Santa Cruz, 1:800).

Dong J., Viswanathan S., Adami E., Singh B.K., Chothani S.P., Ng B., Lim W.W., Zhou J., Tripathi M., Ko N.S., Shekeran S.G., Tan J., Lim S.Y., Wang M., Lio P.M., Yen P.M., Schafer S., Cook S.A, & Widjaja A.A. (2021). Hepatocyte-specific IL11 cis-signaling drives lipotoxicity and underlies the transition from NAFLD to NASH. Nature Communications, 12, 66.

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