Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

Di-Methyl-Histone H3 (Lys9) Antibody #9753

chip   chip validated   chip-on-chip   chipkit   chromatin ip   H3.1   H3.2   H3K9   H3K9me2   histone H3   immunoprecipitation  

No. Size Price
9753L 300 µl ( 30 western blots ) ¥8,792.00 现货查询 购买询价
9753S 100 µl ( 10 western blots ) ¥3,900.00 现货查询 购买询价
9753 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Monkey,D. melanogaster, Endogenous 17 Rabbit
IP 1:50
ChIP 1:25

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, ChIP=Chromatin IP,

Specificity / Sensitivity

Di-Methyl-Histone H3 (Lys9) Antibody 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 Lys9. In addition, the antibody does not cross-react with di-methylated or tri-methylated histone H3 Lys27.

Di-Methyl-Histone H3 (Lys9) Antibody检测仅在Lys9位点双甲基化的内源性histone H3的蛋白水平。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Lys27位点双或三甲基化的histone H3蛋白发生交叉反应。

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the amino terminus of histone H3 in which lysine 9 is di-methylated. Antibodies are purified by protein A and peptide affinity chromatography.

通过合成仅在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多克隆抗体。通过蛋白A和多肽亲和层析纯化获得。

Western Blotting

Western Blotting

Western blot analysis of whole cell lysates from HeLa, NIH/3T3, H-4-II-E and COS cells, using Di-Methyl-Histone H3 (Lys9) Antibody.

使用Di-Methyl-Histone H3 (Lys9) Antibody,免疫印迹(Western blot)分析HeLa、NIH/3T3、H-4-II-E和COS细胞中Di-Methyl-Histone H3 (Lys9)的蛋白水平。

ELISA-Peptide

ELISA-Peptide

Di-Methyl-Histone H3 (Lys9) Antibody 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)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。

IF-IC

IF-IC

Confocal immunofluorescent analysis of NIH/3T3 cells showing nuclear localization when labeled with Di-Methyl-Histone H3 (Lys9) Antibody (green). Actin filaments have been labeled with Alexa Fluor® 555 phalloidin (red).

使用Di-Methyl-Histone H3 (Lys9) Antibody (绿色)标记,共聚焦免疫荧光分析NIH/3T3细胞,结果显示为细胞核定位。Alexa Fluor® 555 phalloidin标记微丝蛋白(红色)。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma, using Di-Methyl-Histone H3 (Lys9) Antibody.

使用Di-Methyl-Histone H3 (Lys9) Antibody,免疫组化分析人源乳腺癌组织石蜡切片。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human Non-Hodgkin''''s lymphoma, using Di-Methyl-Histone H3 (Lys9) Antibody.

使用Di-Methyl-Histone H3 (Lys9) Antibody,免疫组化分析人源非霍奇金淋巴瘤 组织石蜡切片。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human benign prostate hypertrophy (BPH), using Di-Methyl-Histone H3 (Lys9) Antibody.

使用Di-Methyl-Histone H3 (Lys9) Antibody,免疫组化分析人源良性前列腺增生症(BPH)组织石蜡切片。

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) Antibody or 2 μl 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 AFM Intron 1 Primers #5098, SimpleChIP® Human α Satellite Repeat Primers #4486, SimpleChIP® Human RPL30 Exon 3 Primers #7014, and SimpleChIP® Human GAPDH Exon 1 Primers #5516. 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) Antibody或2 µl Normal Rabbit IgG #2729进行染色质免疫沉淀实验。使用SimpleChIP® Human AFM Intron 1 Primers #5098、SimpleChIP® Human α Satellite Repeat Primers #4486、SimpleChIP® Human RPL30 Exon 3 Primers #7014和SimpleChIP® Human GAPDH Exon 1 Primers #5516,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,相当于一个。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lymphoma using Di-Methyl-Histone H3 (Lys9) Antibody in the presence of non-methyl peptide (left) or K9 di-methyl peptide (right).

使用Di-Methyl-Histone H3 (Lys9) (C75H12) Antibody免疫组化分析人源淋巴瘤组织石蜡切片,左图: 非甲基化肽段,右图:K9 双甲基化肽段。

Background

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表遗传标记物(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.

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