Revision 9

#14859Store 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, W-S, IHC-P, IF-F, IF-IC, FC-FP, ChIP, ChIP-seq, C&R

REACTIVITY:

H M

SENSITIVITY:

Endogenous

MW (kDa):

50-70

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q13422

Entrez-Gene Id:

10320

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 5 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.
Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10 - 1:50
Immunohistochemistry (Paraffin) 1:250 - 1:1000
Immunofluorescence (Frozen) 1:50 - 1:100
Immunofluorescence (Immunocytochemistry) 1:100
Flow Cytometry (Fixed/Permeabilized) 1:50 - 1:200
Chromatin IP 1:100
Chromatin IP-seq 1:100
CUT&RUN 1:100

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 #30979.

Specificity / Sensitivity

Ikaros (D6N9Y) Rabbit mAb recognizes endogenous levels of total Ikaros protein.

Species Reactivity:

Human, Mouse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro329 of human Ikaros protein.

Background

The Ikaros family of zinc-finger DNA-binding proteins belongs to the Kruppel transcription factor superfamily. Ikaros proteins are characterized by the presence of an amino-terminal zinc finger DNA-binding domain and a carboxy-terminal dimerization domain. Members of the Ikaros family include Ikaros, Aiolos, Helios, EOS, and Pegasus (1). All family members can form homodimers and heterodimers with other members of the Ikaros family. Most also contain multiple isoforms that are generated as a result of differential splicing, with some isoforms behaving in a dominant negative manner upon dimerization (2).

Ikaros (IKZF1, LYF1) is the prototypical Ikaros family zinc-finger transcription factor and is expressed abundantly in lymphoid cells. Genetic studies in mice demonstrate that Ikaros is a tumor suppressor that is important for the normal development of B, T, natural killer, and dendritic cells (3,4). Additional studies show that imbalanced expression of different Ikaros isoforms, as well as mutations in the corresponding IKAROS gene, can be associated with a number of hematologic malignancies in humans (2,5,6).

  1. Perdomo, J. et al. (2000) J Biol Chem 275, 38347-54.
  2. Rebollo, A. and Schmitt, C. (2003) Immunol Cell Biol 81, 171-5.
  3. Georgopoulos, K. et al. (1997) Annu Rev Immunol 15, 155-76.
  4. Nichogiannopoulou, A. et al. (1998) Semin Immunol 10, 119-25.
  5. Tonnelle, C. et al. (2002) Leuk Lymphoma 43, 29-35.
  6. Martinelli, G. et al. (2009) J Clin Oncol 27, 5202-7.

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 nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting W-S: Simple Western™ IHC-P: Immunohistochemistry (Paraffin) IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq C&R: CUT&RUN

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.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
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 9
#14859

Ikaros (D6N9Y) Rabbit mAb

Western Blotting Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb(上)和 β-Actin (D6A8) Rabbit mAb #8457(下)对不同细胞系的提取物进行蛋白质印迹分析。如预期,可检测的表达完全限于淋巴样细胞系。
Western Blotting Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
Simple Western™ analysis of lysates (0.1 mg/ml) from Jurkat cells using Ikaros (D6N9Y) Rabbit mAb. 虚拟泳道式图像(左图)显示一抗稀释比例在 1:10 和 1:50 时的靶标条带(如图所示)。对应的电泳图(右图)为一抗稀释比例在 1:10(蓝线)和 1:50(绿线)时沿毛细血管内分子量的化学发光结果。This experiment was performed under reducing conditions on the Jess™ Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230kDa.
Immunohistochemistry Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb 对石蜡包埋的人结肠腺癌进行免疫组织化学分析。
Immunohistochemistry Image 2: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb 对石蜡包埋的人非霍奇金淋巴瘤进行免疫组织化学分析。
Immunofluorescence Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb #14859(红色)和 CD11b/ITGAM (M1/70) Rat mAb #46512(绿色)对小鼠皮质进行共聚焦免疫荧光分析。样本用 ProLong® Gold Antifade Reagent with DAPI #8961(蓝色)封片。
Immunofluorescence Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb(绿色)对 A20 细胞(左)、Ramos 细胞(中)和 HeLa 细胞(右)进行共聚焦免疫荧光分析。肌动蛋白微丝由 DyLight 554 Phalloidin #13054(红色)标记。
Flow Cytometry Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 Ikaros (D6N9Y) Rabbit mAb(实线)或浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(虚线)对 HT-29 细胞(蓝色,阴性)和 Jurkat 细胞(绿色,阳性)进行流式细胞分析。Anti-rabbit IgG (H+L)、F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 作为二抗。
Chromatin Immunoprecipitation Image 1: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005,对 Jurkat 细胞中提取的交联染色质和 Ikaros (D6N9Y) Rabbit mAb 进行染色质免疫沉淀。使用 SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795 制成 DNA 库。结果图显示整个 TLR9 基因范围的结合作用。如需了解其他 ChIP-seq 情况,请下载产品说明书。
Chromatin Immunoprecipitation Image 2: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9005,对 Jurkat 细胞中提取的交联染色质和 Ikaros (D6N9Y) Rabbit mAb 进行染色质免疫沉淀。使用 SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795 制成 DNA 库。结果图显示在染色体 3(上图)内的结合,包括 TLR9 基因(下图)。
Chromatin Immunoprecipitation Image 3: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003,对 Jurkat 细胞中提取的交联染色质,在加入 Ikaros (D6N9Y) Rabbit mAb 或 μl Normal Rabbit IgG #2729 后,进行染色质免疫沉淀。使用人 HES1 启动子引物、SimpleChIP® Human HES4 Promoter Primers #7273 和 SimpleChIP® Human α Satellite Repeat Primers #4486,通过实时 PCR 对富集的 DNA 进行定量。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: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 Jurkat 细胞和 Ikaros (D6N9Y) Rabbit mAb 进行 CUT&RUN 检测。‚使用 SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795 制备 DNA 库。该图显示在 HES1 基因内结合。
CUT and RUN Image 2: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 Jurkat 细胞和 Ikaros (D6N9Y) Rabbit mAb 进行 CUT&RUN 检测。‚使用 SimpleChIP® ChIP-seq DNA Library Prep Kit for Illumina® #56795 制备 DNA 库。该图显示在染色体 3(上图)内的结合,包括 HES1 基因(下图)。
CUT and RUN Image 3: Ikaros (D6N9Y) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652 对 Jurkat 细胞和 Ikaros (D6N9Y) Rabbit mAb 或 Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362 进行 CUT&RUN 检测。使用 human TMEM exon 1、human CKS1B exon 1 和 SimpleChIP® Human α Satellite Repeat Primers #4486 进行实时 PCR 来对富集的 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。