Cell Signaling Technology

Product Pathways - Chromatin Regulation / Epigenetics

Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb #8647

sc-377521  

No. Size Price
8647S 100 µl ( 10 western blots ) ¥3,900.00 现货查询 购买询价
8647 carrier free & custom formulation / quantityemail request
Applications Dilution Species-Reactivity Sensitivity MW (kDa) Isotype
W 1:1000 Human,Mouse,Rat,Monkey, Endogenous 11 Rabbit IgG
IP 1:100
IHC-P 1:6400
IF-IC 1:100
ChIP 1:25

Species cross-reactivity is determined by western blot.

Applications Key: W=Western Blotting, IP=Immunoprecipitation, IHC-P=Immunohistochemistry (Paraffin), IF-IC=Immunofluorescence (Immunocytochemistry), ChIP=Chromatin IP,

Homology

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

Specificity / Sensitivity

Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb recognizes endogenous levels of histone H4 protein only when acetylated at Lys5. This antibody does not cross-react with histone H4 acetylated at Lys8, Lys12, or Lys16.

Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb兔单抗能够检测仅在Lys5位点乙酰化的内源性histone H4蛋白水平。该抗体不能与lysines 8、12或16位点乙酰化的histone H4发生交叉反应。

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding acetylated Lys5 of human histone H4 protein.

通过人工合成人源histone H4蛋白Lys5位点乙酰化的周围相应的片段去免疫动物从而制备出此多克隆抗体。

IF-IC

IF-IC

Confocal immunofluorescent analysis of HeLa cells, untreated (left) or treated with Trichostatin A (TSA) #9950 (right) using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb (green). Actin filaments were labeled with DY-554 phalloidin (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb (绿色),共聚焦免疫荧光分析untreated (左图)或treated Trichostatin A (TSA) #9950 (右图)的HeLa细胞。DY-554 phalloidin标记微丝蛋白(红色)。蓝色= DRAQ5® #4084 (DNA荧光染料)。

Chromatin IP

Chromatin IP

Chromatin immunoprecipitations were performed with cross-linked chromatin from 4 x 106 HeLa cells and either 20 µl of Acetyl-Histone H4 (Lys5) (D12B3) 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 MyoD1 Exon 1 Primers #4490, 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,来自4 x 106 HeLa细胞的交联染色质以及20 µl Acetyl-Histone H4 (Lys5) (D12B3) 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 MyoD1 Exon 1 Primers #4490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1的总input chromatin的数量的信号。

Western Blotting

Western Blotting

Western blot analysis of extracts from various cell lines using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb.

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb兔单抗,免疫印迹(Western blot)分析不同细胞中Acetyl-Histone H4 (Lys5) 蛋白水平。

Western Blotting

Western Blotting

Western blot analysis of extracts from HeLa and C2C12 cells, untreated (-) or treated with Trichostatin A (TSA) #9950 (1 μM, 18 hr; +), using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb (upper) or Histone H4 (L64C1) Mouse mAb #2935 (lower).

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb (上图)或Histone H4 (L64C1) Mouse mAb #2935 (下图),免疫印迹(Western blot)分析不同细胞中Acetyl-Histone H4 (Lys5)和Histone H4蛋白水平,细胞分为未处理组 (-)或处理组: Trichostatin A (TSA) #9950 (1 μM, 18 hr; +)。

ELISA-Peptide

ELISA-Peptide

Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb specificity was determined by peptide ELISA. The graph depicts the binding of the antibody to pre-coated acetyl-histone H4 (Lys5) peptide in the presence of increasing concentrations of various competitor peptides. As shown, only the acetyl-histone H4 (Lys5) peptide competed away binding of the antibody.

通过peptide ELISA确定Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb的特异性。该图描述了抗体与提前包被的acetyl-histone H4 (Lys5) 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acetyl-histone H4 (Lys5) peptide竞争脱离抗体的结合。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb in the presence of non-acetyl-histone H4 (Lys5) peptide (left) or acetyl-histone H4 (Lys5) peptide (right).

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb,该抗体与non-acetyl-histone H4 (Lys5) peptide (左图)或acetyl-histone H4 (Lys5) peptide (右图)一起孵育,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 colon carcinoma using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb.

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb兔单抗,免疫组化分析人源结肠癌组织石蜡切片。

IHC-P (paraffin)

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb.

使用Acetyl-Histone H4 (Lys5) (D12B3) Rabbit mAb兔单抗,免疫组化分析人源肺癌组织石蜡切片。

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 (1,2). Histone acetylation occurs mainly on the amino-terminal tail domains of histones H2A (Lys5), H2B (Lys5, 12, 15, and 20), H3 (Lys9, 14, 18, 23, 27, and 56), and H4 (Lys5, 8, 12, and 16) and is important for the regulation of histone deposition, transcriptional activation, DNA replication, recombination, and DNA repair (1-3). Hyper-acetylation of the histone tails neutralizes the positive charge of these domains and is believed to weaken histone-DNA and nucleosome-nucleosome interactions, thereby destabilizing chromatin structure and increasing the accessibility of DNA to various DNA-binding proteins (4,5). In addition, acetylation of specific lysine residues creates docking sites for a protein module called the bromodomain, which binds to acetylated lysine residues (6). Many transcription and chromatin regulatory proteins contain bromodomains and may be recruited to gene promoters, in part, through binding of acetylated histone tails. Histone acetylation is mediated by histone acetyltransferases (HATs), such as CBP/p300, GCN5L2, PCAF, and Tip60, which are recruited to genes by DNA-bound protein factors to facilitate transcriptional activation (3). Deacetylation, which is mediated by histone deacetylases (HDAC and sirtuin proteins), reverses the effects of acetylation and generally facilitates transcriptional repression (7,8).

核小体是由四种中心组蛋白(H2A、H2B、H3和H4)组成,它是染色质的主要构件模块。起初被认为是一个DNA包装的静态支架,目前组蛋白已经被认为是一个动态蛋白,其经历多种形式的翻译后修饰,包括乙酰化作用、磷酸化作用、甲基化作用和泛素化作用(1,2)。组蛋白乙酰化作用主要发生在histones H2A (Lys5)、H2B (Lys5, 12, 15, and 20)、H3 (Lys9, 14, 18, 23, 27, and 56)和H4 (Lys5, 8, 12, and 16)的氨基端尾部结构域,并且对于组蛋白沉积、转录激活、DNA复制、重组和DNA修复有着重要作用(1-3)。组蛋白尾部的高度乙酰化中和了这些区域的正电荷,并且被认为弱化了组蛋白-DNA和核小体-核小体的相互作用,因此使核染色质不稳定以及增加了DNA与不同的DNA结合蛋白的接近程度(4,5)。此外,特定的赖氨酸残基乙酰化产生了停留点,这些位置有助于招募包含一个bromodomain蛋白,该区域能结合到乙酰化的赖氨酸残基(6)。许多转录和染色质调节蛋白包含bromodomains,并且可能被招募到基因启动子,在某种程度上通过乙酰化的组蛋白尾部结合。组蛋白乙酰化通过组蛋白乙酰化转移酶(HATs)介导,例如CBP/p300、GCN5L2、PCAF和Tip60,这些通过DNA结合蛋白因子招募到基因上从而有助于转录激活(3)。去乙酰化作用通过组蛋白去乙酰转移酶(HDAC和Sirtuin蛋白)介导,这将逆转乙酰化的影响,并且有助于转录抑制(7,8)。

  1. Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
  2. Jaskelioff, M. and Peterson, C.L. (2003) Nat Cell Biol 5, 395-9.
  3. Roth, S.Y. et al. (2001) Annu Rev Biochem 70, 81-120.
  4. Workman, J.L. and Kingston, R.E. (1998) Annu Rev Biochem 67, 545-79.
  5. Hansen, J.C. et al. (1998) Biochemistry 37, 17637-41.
  6. Yang, X.J. (2004) Bioessays 26, 1076-87.
  7. Haberland, M. et al. (2009) Nat Rev Genet 10, 32-42.
  8. Haigis, M.C. and Sinclair, D.A. (2010) Annu Rev Pathol 5, 253-95.

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