Revision 7

#14696Store 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, ChIP, ChIP-seq, C&R, C&T

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

43, 48

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q9HC52

Entrez-Gene Id:

57332

Product Information

Product Usage Information

For optimal ChIP and 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 1:200
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.

Specificity / Sensitivity

CBX8 (D2O8C) Rabbit mAb recognizes endogenous levels of total CBX8 protein.

Species Reactivity:

Human, Mouse, Rat, Monkey

Species predicted to react based on 100% sequence homology

Hamster, Bovine

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Gly344 of human CBX8 protein.

Background

The polycomb group (PcG) proteins contribute to the maintenance of cell identity, stem cell self-renewal, cell cycle regulation and oncogenesis by maintaining the silenced state of genes that promote cell lineage specification, cell death and cell-cycle arrest (1-4). PcG proteins exist in two complexes that cooperate to maintain long-term gene silencing through epigenetic chromatin modifications: PRC1 and PRC2. PRC1 is a multi-subunit protein complex consisting of a combination of five core protein families: CBX, RING1, PHC, PCGF, and RYPB (5-7). Different combinations of protein family members lead to a diverse array of PRC1 complexes with distinct functions (8). At least two distinct classes of PRC1 complexes have been defined. The first class, known as canonical PRC1, contains RING1, PHC, PCGF and CBX protein subunits, but not RYPB (5-8). This class of PRC1 complexes requires PRC2 and H3K27Me3 for proper recruitment to target genes. CBX proteins mediate recruitment by binding to H3K27Me3. CBX8 in particular is required for repression of many lineage-specific genes during differentiation of hematopoietic stem cells and may play a role in activation of lineage-specific genes during differentiation of embryonic stem cells (9,10). The second class, known as variant PRC1, contains RYPB instead of CBX proteins (5-8). RYBP-containing PRC1 is recruited to chromatin independently of PRC2 and H3K27Me3. These variant PRC1 complexes can function independently of PRC2, or in some cases function upstream to recruit PRC2 complex to target genes.

  1. Boyer, L.A. et al. (2006) Nature 441, 349-53.
  2. Lee, T.I. et al. (2006) Cell 125, 301-13.
  3. Park, I.K. et al. (2003) Nature 423, 302-5.
  4. Molofsky, A.V. et al. (2003) Nature 425, 962-7.
  5. Tavares, L. et al. (2012) Cell 148, 664-78.
  6. Gao, Z. et al. (2012) Mol Cell 45, 344-56.
  7. Blackledge, N.P. et al. (2014) Cell 157, 1445-59.
  8. Luis, N.M. et al. (2012) Cell Stem Cell 11, 16-21.
  9. Klauke, K. et al. (2013) Nat Cell Biol 15, 353-62.
  10. Creppe, C. et al. (2014) PLoS Genet 10, e1004851.

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 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.
SimpleChIP is a registered 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 7
#14696

CBX8 (D2O8C) Rabbit mAb

Western Blotting Image 1: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 CBX8 (D2O8C) Rabbit mAb,对不同细胞系提取物进行蛋白质印迹法分析。
Immunoprecipitation Image 1: CBX8 (D2O8C) Rabbit mAb Expand Image
对 A549 细胞提取物的 CBX8 进行免疫沉淀。泳道 1 是 10% 输入,泳道 2 是 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900,泳道 3 是 CBX8 (D2O8C) Rabbit mAb。使用 CBX8 (D2O8C) Rabbit mAb,进行蛋白质印迹法分析。
Chromatin Immunoprecipitation Image 1: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005 对 NCCIT 细胞中提取的交联染色质,在加入 JARID2 (D6M9X) Rabbit mAb#13594, CBX8 (D2O8C) Rabbit mAb 或 AEBP2 (D7C6X) Rabbit mAb #14129 后,进行染色质免疫沉淀法。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。所有 JARID2、CBX8 和 AEBP2 已知可结合 PRC 复合体。本图显示整个 HOXD 基因簇范围内 JARID2, CBX8 和 AEBP2(其为 JARID2、CBX8 和 AEBP2 的已知靶标基因)的结合作用(参见包含 ChIP-qPCR 数据的其他结果图)。
Chromatin Immunoprecipitation Image 2: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005 对 NCCIT 细胞中提取的交联染色质,在加入 JARID2 (D6M9X) Rabbit mAb#13594, CBX8 (D2O8C) Rabbit mAb 或 AEBP2 (D7C6X) Rabbit mAb #14129 后,进行染色质免疫沉淀法。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。所有 JARID2、CBX8 和 AEBP2 已知可结合 PRC 复合体。本图显示整个 HOXA(上图)和 HOXD(下图)范围内 JARID2、CBX8 和 AEBP2(其为 JARID2、CBX8 和 AEBP2 的已知靶标基因)的结合作用(参见包含 ChIP-qPCR 数据的其他结果图)。
Chromatin Immunoprecipitation Image 3: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,对 NCCIT 细胞中提取的交联染色质,在加入 CBX8 (D2O8C) Rabbit mAb 或Normal Rabbit IgG #2729,进行染色质免疫沉淀。使用 SimpleChIP® Human HoxA1 Intron 1 Primers #7707、SimpleChIP® Human HoxA2 Promoter Primers #5517 和 SimpleChIP® Human α Satellite Repeat Primers #4486 通过实时 PCR 对富集 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: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 NCCIT 细胞和 CBX8 (D2O8C) Rabbit mAb 进行 CUT&RUN 分析。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。该图显示在 HOXA1 内结合,HOXA1 是 CBX8 的已知靶标基因(参见包含 qPCR 数据的其他结果图)。
CUT and RUN Image 2: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 NCCIT 细胞和 CBX8 (D2O8C) Rabbit mAb 进行 CUT&RUN 分析。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。这些图显示了 CBX8 的已知靶标基因 HoxD(上图)和 HoxA(下图)之间的结合(参见包含 qPCR 数据的其他结果图)。
CUT and RUN Image 3: CBX8 (D2O8C) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 NCCIT 和 CBX8 (D2O8C) Rabbit mAb 或 Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362 进行 CUT&RUN。使用 SimpleChIP® Human HoxA1 Intron 1 Primers #7707、SimpleChIP® Human HoxA2 Promoter Primers #5517 和 SimpleChIP® Human α Satellite Repeat Primers #4486 通过实时 PCR 对富集 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: CBX8 (D2O8C) Rabbit mAb Expand Image
CUT&Tag was performed with NCCIT cells and CBX8 (D2O8C) 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 HOXA1, a known target gene of CBX8 (see ChIP-qPCR figure).
CUT and Tag Image 2: CBX8 (D2O8C) Rabbit mAb Expand Image
CUT&Tag was performed with NCCIT cells and CBX8 (D2O8C) 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 the HOXA gene cluster (upper), and the HOXD gene cluster (lower), which are known target genes of CBX8 (see our ChIP-qPCR figure).