Revision 8

#3418Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IHC-P, IF-F, IF-IC, ChIP, ChIP-seq, C&R, C&T

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

140

Source/Isotype:

Rabbit IgG

UniProt ID:

#P49711

Entrez-Gene Id:

10664

Product Information

Product Usage Information

For optimal ChIP or ChIP-seq results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

The CUT&RUN dilution was determined using CUT&RUN Assay Kit #86652.

The CUT&Tag dilution was determined using CUT&Tag Assay Kit #77552.

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunohistochemistry (Paraffin) 1:640 - 1:2560
Immunofluorescence (Frozen) 1:200 - 1:800
Immunofluorescence (Immunocytochemistry) 1:400 - 1:1600
Chromatin IP 1:50
Chromatin IP-seq 1:50
CUT&RUN 1:50
CUT&Tag 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #48859.

Specificity / Sensitivity

CTCF (D31H2) XP® Rabbit mAb detects endogenous levels of total CTCF protein. This antibody does not cross-react with BORIS. This antibody nonspecifically labels the lamina propria of small intestine in fixed frozen mouse tissue by immunofluorescence.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Bovine, Horse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to the carboxy terminus of the human CTCF protein.

Background

CCCTC-binding factor (CTCF) and its paralog, the Brother of the Regulator of Imprinted Sites (BORIS), are highly conserved transcription factors that regulate transcriptional activation and repression, insulator function, and imprinting control regions (ICRs) (1-4). Although they have divergent amino and carboxy termini, both proteins contain 11 conserved zinc finger domains that work in combination to bind the same DNA elements (1). CTCF is ubiquitously expressed and contributes to transcriptional regulation of cell-growth regulated genes, including c-Myc, p19/ARF, p16/INK4A, BRCA1, p53, p27, E2F1, and TERT (1). CTCF also binds to and is required for the enhancer-blocking activity of all known insulator elements and ICRs, including the H19/IgF2, Prader-Willi/Angelman syndrome, and Inactive X-Specific Transcript (XIST) anti-sense loci (5-7). CTCF DNA-binding is sensitive to DNA methylation, a mark that determines selection of the imprinted allele (maternal vs. paternal) (1). The various functions of CTCF are regulated by at least two different post-translational modifications. Poly(ADP-ribosyl)ation of CTCF is required for insulator function (8). Phosphorylation of Ser612 by protein kinase CK2 facilitates a switch of CTCF from a transcriptional repressor to an activator at the c-Myc promoter (9). CTCF mutations or deletions have been found in many breast, prostate, and Wilms' tumors (10,11). Expression of BORIS is restricted to spermatocytes and is mutually exclusive of CTCF (3). In cells expressing BORIS, promoters of X-linked cancer-testis antigens like MAGE-A1 are demethylated and activated, but methylated and inactive in CTCF-expressing somatic cells (12). Like other testis specific proteins, BORIS is abnormally expressed in different cancers, such as breast cancer, and has a greater affinity than CTCF for DNA-binding sites, detracting from CTCF’s potential tumor suppressing activity (1,3,13,14).

  1. Klenova, E.M. et al. (2002) Semin Cancer Biol 12, 399-414.
  2. Klenova, E.M. et al. (1993) Mol Cell Biol 13, 7612-24.
  3. Loukinov, D.I. et al. (2002) Proc Natl Acad Sci USA 99, 6806-11.
  4. Mukhopadhyay, R. et al. (2004) Genome Res 14, 1594-602.
  5. Hark, A.T. et al. (2000) Nature 405, 486-9.
  6. Ohta, T. et al. (1999) Am J Hum Genet 64, 397-413.
  7. Chao, W. et al. (2002) Science 295, 345-7.
  8. Yu, W. et al. (2004) Nat Genet 36, 1105-10.
  9. El-Kady, A. and Klenova, E. (2005) FEBS Lett 579, 1424-34.
  10. Filippova, G.N. et al. (1998) Genes Chromosomes Cancer 22, 26-36.
  11. Filippova, G.N. et al. (2002) Cancer Res 62, 48-52.
  12. Vatolin, S. et al. (2005) Cancer Res 65, 7751-62.
  13. Hong, J.A. et al. (2005) Cancer Res 65, 7763-74.
  14. D'Arcy, V. et al. (2008) Br J Cancer 98, 571-9.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IHC-P: Immunohistochemistry (Paraffin) IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq C&R: CUT&RUN C&T: CUT&Tag

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意,否书以下条款适用于 CST、其关书方或分书商提供的书品。 任何书充本条款或与本条款不同的客书条款和条件,除非书 CST 的合法授书代表以书面形式书独接受, 否书均被拒书,并且无效。

专品专有“专供研究使用”的专专或专似的专专声明, 且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的, 或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专,且专用于研专用途。将专品用于专断、专防或治专目的, 或专专售(专独或作专专成)或其他商专目的而专专专品,均需要 CST 的专独专可。客专:(a) 不得专独或与其他材料专合向任何第三方出售、专可、 出借、捐专或以其他方式专专或提供任何专品,或使用专品制造任何商专专品,(b) 不得复制、修改、逆向工程、反专专、 反专专专品或以其他方式专专专专专品的基专专专或技专,或使用专品开专任何与 CST 的专品或服专专争的专品或服专, (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专,(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品, (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Revision 8
#3418

CTCF (D31H2) XP® Rabbit mAb

Western Blotting Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CTCF (D31H2) XP® Rabbit mAb 对不同细胞系提取物进行蛋白质印迹分析。
No image available
Immunohistochemistry Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
在对照肽(左)或抗原特异性肽(右)存在的情况下,使用 CTCF (D31H2) XP® Rabbit mAb 对石蜡包埋的人乳腺癌进行免疫组织化学分析。
Immunohistochemistry Image 2: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CTCF (D31H2) XP® Rabbit mAb 对石蜡包埋的人结肠癌进行免疫组织化学分析。
Immunohistochemistry Image 3: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CTCF (D31H2) XP® Rabbit mAb 对石蜡包埋的人淋巴瘤进行免疫组织化学分析。
Immunofluorescence Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CTCF (D31H2) XP® Rabbit mAb(绿色)对 HCT-116 细胞进行共聚焦免疫荧光分析。使用 DY-554 phalloidin(红色)标记肌动蛋白纤丝。
Chromatin Immunoprecipitation Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,对 Hela 细胞中提取的交联染色质,在加入 CTCF (D31H2) XP® Rabbit mAb 后进行染色质免疫沉淀。使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。图形显示在 H19 内结合,H19 是一种已知的 CTCF 靶标基因(参见包含 ChIP-qPCR 数据的其他图形)。如需了解其他 ChIP-seq 情况,请下载产品说明书。
Chromatin Immunoprecipitation Image 2: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,对 Hela 细胞中提取的交联染色质,在加入 CTCF (D31H2) XP® Rabbit mAb 后进行染色质免疫沉淀。使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。结果图显示在染色体 11 内的结合(上图),包括 CTCF 的已知靶基因 H19(下图)(参见包含 ChIP-qPCR 数据的其他结果图)。
Chromatin Immunoprecipitation Image 3: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,对 HeLa 细胞中提取的交联染色质,在加入 CTCF (D31H2) XP® Rabbit mAb 或 Normal Rabbit IgG #2729 后进行染色质免疫沉淀。使用 Human c-Myc Promoter Primers、SimpleChIP® Human H19/Igf2 ICR Primers #5172 和 SimpleChIP® Human α Satellite Repeat Primers #4486 并通过实时 PCR 对富集的 DNA 进行定量分析。将每份样品中免疫沉淀的 DNA 的量表现为相对于所输入染色质总量(等于 1)的信号。
CUT and RUN Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652对 HCT 116 细胞和 CTCF (D31H2) XP® Rabbit mAb #3418 进行 CUT&RUN。使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。图形显示在 MYC 内结合,MYC 是一种已知的 CTCF 靶标基因(参见包含 CUT&RUN-qPCR 数据的其他图形)。
CUT and RUN Image 2: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652对 HCT 116 细胞和 CTCF (D31H2) XP® Rabbit mAb #3418 进行 CUT&RUN。使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。结果图显示在染色体 8(上图)内的结合,包括 CTCF 的已知靶基因 MYC(下图)(参见包含 CUT&RUN-qPCR 数据的其他结果图)。
CUT and RUN Image 3: CTCF (D31H2) XP® Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对HCT 116 细胞和 CTCF (D31H2) XP® Rabbit mAb 或 Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT & RUN) #66362 进行 CUT&RUN。使用 Human c-Myc Promoter Primers、SimpleChIP® Human H19/Igf2 ICR Primers #5172 和 SimpleChIP® Human α Satellite Repeat Primers #4486 并通过实时 PCR 对富集的 DNA 进行定量分析。将每份样品中免疫沉淀的 DNA 的量表现为相对于所输入染色质总量(等于 1)的信号。
CUT and Tag Image 1: CTCF (D31H2) XP® Rabbit mAb Expand Image
CUT&Tag was performed with HCT 116 cells and CTCF (D31H2) XP® Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figure shows binding across MYC, a known target gene of CTCF (see our ChIP-qPCR figure).
CUT and Tag Image 2: CTCF (D31H2) XP® Rabbit mAb Expand Image
CUT&Tag was performed with HCT 116 cells and CTCF (D31H2) XP® Rabbit mAb, using CUT&Tag Assay Kit #77552. DNA library was prepared using CUT&Tag Dual Index Primers and PCR Master Mix for Illumina Systems #47415. The figures show binding across chromosome 8 (upper), including MYC (lower), a known target gene of CTCF (see our ChIP-qPCR figure).