Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

Di-Methyl-Histone H3 (Lys9) (D85B4) XP® Rabbit mAb #4658

H3.1   H3.2   H3K9   H3K9me2   Histone   methyl   methylation  

No. Size Price
4658S 100 µl ( 10 western blots ) ¥4,180.00 现货查询 购买询价 防伪查询
4658T 20 µl ( 2 western blots ) ¥1,600.00 现货查询 购买询价 防伪查询
4658 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Monkey, Endogenous 17 Rabbit IgG
IP 1:50
F 1:100
IF-IC 1:200
ChIP 1:50

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, F=Flow Cytometry, IF-IC=Immunofluorescence (Immunocytochemistry), ChIP=Chromatin IP,


Species predicted to react based on 100% sequence homology: D. melanogaster, Xenopus, Zebrafish, Bovine, Pig, S. cerevisiae, C. elegans,

Specificity / Sensitivity

Di-Methyl-Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb detects endogenous levels of histone H3 only when di-methylated on Lys9. The antibody does not cross-react with non-methylated, mono-methylated or tri-methylated Histone H3 Lys9. In addition, the antibody does not cross-react with histone H3 di-methylated on Lys4, Lys27, or Lys36 and histone H4 di-methylated on Lys20.

Di-Methyl-Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb兔单抗检测仅在Lys9位点双甲基化的内源性histone H3。该抗体不与Lys9位点非甲基化、单甲基化或三甲基化的histone H3发生交叉反应。另外,该抗体不与Lys4、Lys9、Lys27、Lys36位点甲基化的histone H3蛋白或Lys20位点甲基化的histone H4蛋白发生交叉反应。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to the amino terminus of histone H3 in which Lys9 is di-methylated.

通过合成的仅在lysine 9位点双甲基化histone H3蛋白氨基端相应的多肽片段去免疫动物从而制备出此单克隆抗体。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Di-Methyl-Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb.

使用Di-Methyl-Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb兔单抗,免疫印迹(Western blot)分析不同细胞中Di-Methyl-Histone H3 (Lys9)蛋白水平。



Di-Methyl Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb specificity was determined by peptide ELISA. The graph depicts the binding of the antibody to pre-coated di-methyl histone H3 (Lys9) peptide in the presence of increasing concentrations of various competitor peptides. As shown, only the di-methyl histone H3 (Lys9) peptide competed away binding of the antibody.

通过peptide ELISA确定Di-Methyl Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb的特异性。该图描述了抗体与提前包被的di-methyl histone H3 (Lys9) 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 (Lys9) peptide竞争脱离抗体的结合。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 20 μl of Di-Methyl-Histone H3 (Lys9) (D85B4) 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 RPL30 Exon 3 Primers #7014, SimpleChIP® Human AFM Intron 1 Primers #5098, and SimpleChIP® Human α Satellite Repeat Primers #4486. 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20 µl Di-Methyl-Histone H3 (Lys9) (D85B4) 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 RPL30 Exon 3 Primers #7014、SimpleChIP® Human AFM Intron 1 Primers #5098和SimpleChIP® Human α Satellite Repeat Primers #4486,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,相当于一。



Confocal immunofluorescent analysis of HeLa cells using Di-Methyl-Histone H3 (Lys9) (D85B4) XP™ Rabbit mAb(green). Actin filaments have been labeled with DY-554 phalloidin (red).

使用Di-Methyl-Histone H3 (Lys9) (D85B4) 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)。

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of HeLa cells using Di-Methyl-Histone H3 (Lys9) (D85B4) XP Rabbit mAb (blue) compared to concentration-matched Rabbit (DA1E) mAb IgG XP Isotype Control #3900 (red). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor 488 Conjugate) #4412 was used as a secondary antibody.


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).

核小体是由四种组蛋白(H2A、H2B、H3和H4)组成,它是染色质的主要构成模块。起初被认为作为一个DNA包装的静态支架,现在则显示组蛋白是动态蛋白,经历多种翻译后修饰的形式,包括乙酰化、磷酸化、甲基化和泛素化(1)。组蛋白甲基化对于该基因组的活化和未活化区域的形成有着主要决定作用,并且在发育期间对该基因组的正确规划起着关键作用(2,3)。histones H3 (Arg2、17、26)和H4 (Arg3)的精氨酸甲基化促进转录调控以及通过蛋白质精氨酸甲基转移酶(PRMTs)家族的介导,包括共激活因子PRMT1和CARM1 (PRMT4) (4)。相反,多种多样的组蛋白赖氨酸甲基转移酶已经被鉴定,除了这个之外其它的都包含一个保守的催化SET区域,这个起初被鉴定在Drosophila Su(var)3-9、zeste增强子和Trithorax蛋白。赖氨酸甲基化主要发生在histones H3 (Lys4、9、27、36、79)和H4 (Lys20),并且已经涉及到转录激活和沉默(4)。这些赖氨酸残基的甲基化协调染色质修饰酶的招募包括methyl-lysine结合模块例如chromodomains (HP1, PRC1)、PHD fingers (BPTF, ING2)、tudor domains (53BP1)和WD-40 domains (WDR5) (5-8)。组蛋白例如PADI4、LSD1、JMJD1、JMJD2和JHDM1的发现已经显示甲基化是一个可逆的后生标记物(9)。

  1. Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
  2. Kubicek, S. et al. (2006) Ernst Schering Res Found Workshop , 1-27.
  3. Lin, W. and Dent, S.Y. (2006) Curr Opin Genet Dev 16, 137-42.
  4. Lee, D.Y. et al. (2005) Endocr Rev 26, 147-70.
  5. Daniel, J.A. et al. (2005) Cell Cycle 4, 919-26.
  6. Shi, X. et al. (2006) Nature 442, 96-9.
  7. Wysocka, J. et al. (2006) Nature 442, 86-90.
  8. Wysocka, J. et al. (2005) Cell 121, 859-72.
  9. Trojer, P. and Reinberg, D. (2006) Cell 125, 213-7.

Application References

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