| 货号 | 9728S |
| 反应种属 | Human/Mouse/Rat/Monkey |
| 来源宿主 | Rabbit |
| 应用 | W/IP/IF-IC/F/ChIP |
| 目标/特异性 | Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb detects endogenous levels of histone H3 only when di-methylated on Lys27. The antibody does not cross-react with histone H3 that is non-methylated, mono-methylated or tri-methylated at Lys27. In addition, the antibody does not cross-react with mono-methylated, di-methylated or tri-methylated histone H3 Lys4, Lys9, Lys36 or histone H4 Lys20. |
| 使用方法 | WB(1:1000) IP (1:50) F (1:400) IF-IC (1:3200) ChIP (1:50) |
| 供应商 | CST |
| 背景 | The nucleosome, made up of four core histone proteins (H2A, H2B, H3 and H4), is the primary building block of chromatin. Originally thought to function as a static scaffold for DNA packaging, histones have now been shown to be dynamic proteins, undergoing multiple types of post-translational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (1). Histone methylation is a major determinant for the formation of active and inactive regions of the genome and is crucial for the proper programming of the genome during development (2,3). Arginine methylation of histones H3 (Arg2, 17, 26) and H4 (Arg3) promotes transcriptional activation and is mediated by a family of protein arginine methyltransferases (PRMTs), including the co-activators PRMT1 and CARM1 (PRMT4) (4). In contrast, a more diverse set of histone lysine methyltransferases has been identified, all but one of which contain a conserved catalytic SET domain originally identified in the Drosophila Su(var)3-9, Enhancer of zeste, and Trithorax proteins. Lysine methylation occurs primarily on histones H3 (Lys4, 9, 27, 36, 79) and H4 (Lys20) and has been implicated in both transcriptional activation and silencing (4). Methylation of these lysine residues coordinates the recruitment of chromatin modifying enzymes containing methyl-lysine binding modules such as chromodomains (HP1, PRC1), PHD fingers (BPTF, ING2), tudor domains (53BP1), and WD-40 domains (WDR5) (5-8). The discovery of histone demethylases such as PADI4, LSD1, JMJD1, JMJD2, and JHDM1 has shown that methylation is a reversible epigenetic marker (9). |
| 存放说明 | -20C |
| 计算分子量 | 17 |
Confocal immumunofluorescent analysis of HeLa cells using Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb (green). Actin filaments have been labled with DY-554 phalloidin (red). 使用Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb 兔单抗(绿色)，共聚焦免疫荧光分析HeLa细胞。DY-554 phalloidin标记微丝蛋白(红色)。 | |
Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb specificity was determined by peptide ELISA. The graph depicts the binding of the antibody to pre-coated di-methyl histone H3 (Lys27) peptide in the presence of increasing concentrations of various competitor peptides. As shown, only the di-methyl histone H3 (Lys27) peptide competed away binding of the antibody. 通过peptide ELISA确定Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb兔单抗的特异性。该图描述了抗体与提前包被的di-methyl histone H3 (Lys27) peptide的结合能力，并且多肽中含有浓度递增的不同竞争多肽。如同所示，仅di-methyl histone H3 (Lys27) peptide竞争脱离抗体的结合。 | |
Chromatin immunoprecipitations were performed with cross-linked chromatin from 4x106 HeLa cells and either 10 μl of Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb or 2 μl of Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR, using SimpleChIP® Human GAPDH Exon 1 Primers #5516, SimpleChIP® Human MyoD1 Exon 1 Primers #4490, and SimpleChIP® Human AFM Intron 1 Primers #5098. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one. 使用SimpleChIP®Enzymatic Chromatin IP Kit (Magnetic Beads) #9003，用4 x 106 HeLa细胞的交联染色质以及10 µl Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb或2 µl Normal Rabbit IgG #2729进行染色质免疫沉淀实验。使用SimpleChIP® Human GAPDH Exon 1 Primers #5516、SimpleChIP® Human MyoD1 Exon 1 Primers #4490和SimpleChIP® Human AFM Intron 1 Primers #5098，浓缩的DNA通过real-time PCR定量。在每个样品中免疫沉淀DNA的数量被当做一个相对于总input chromatin的数量的信号，这相当于一。 | |
Western blot analysis of extracts from various cell lines using Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb. 使用Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb兔单抗，免疫印迹(Western blot)分析不同细胞中Di-Methyl-Histone H3 (Lys27)的蛋白水平。 | |
Flow cytometric analysis of human peripheral blood lymphocytes using Di-Methyl-Histone H3 (Lys27) (D18C8) XP® Rabbit mAb (blue) compared to Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab)2 Fragment (Alexa Fluor® 647 Conjugate) #4414 was used as a secondary antibody. |
