Revision 7

#13987Store 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, IF-IC, FC-FP, ChIP, ChIP-seq, C&R

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

57-65

Source/Isotype:

Rabbit IgG

UniProt ID:

#P04198, #P01106

Entrez-Gene Id:

4613, 4609

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.
Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:1600
Flow Cytometry (Fixed/Permeabilized) 1:400
Chromatin IP 1:50
Chromatin IP-seq 1:50
CUT&RUN 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 #83277.

Specificity / Sensitivity

c-Myc/N-Myc (D3N8F) Rabbit mAb recognizes endogenous levels of total c-Myc and N-Myc proteins.

Species Reactivity:

Human, Mouse, Rat, Monkey

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein specific to a central region within human c-Myc protein.

Background

Members of the Myc/Max/Mad network function as transcriptional regulators with roles in various aspects of cell behavior, including proliferation, differentiation, and apoptosis (1). These proteins share a common basic-helix-loop-helix leucine zipper (bHLH-ZIP) motif required for dimerization and DNA-binding. Max was originally discovered based on its ability to associate with c-Myc and found to be required for the ability of Myc to bind DNA and activate transcription (2). Subsequently, Max has been viewed as a central component of the transcriptional network, forming homodimers as well as heterodimers with other members of the Myc and Mad families (1). The association between Max and either Myc or Mad can have opposing effects on transcriptional regulation and cell behavior (1). The Mad family consists of four related proteins; Mad1, Mad2 (Mxi1), Mad3, and Mad4, and the more distantly related members of the bHLH-ZIP family, Mnt and Mga. Like Myc, the Mad proteins are tightly regulated with short half-lives. In general, Mad family members interfere with Myc-mediated processes, such as proliferation, transformation, and prevention of apoptosis by inhibiting transcription (3,4).

  1. Baudino, T.A. and Cleveland, J.L. (2001) Mol Cell Biol 21, 691-702.
  2. Blackwood, E.M. and Eisenman, R.N. (1991) Science 251, 1211-7.
  3. Henriksson, M. and Lüscher, B. (1996) Adv Cancer Res 68, 109-82.
  4. Grandori, C. et al. (2000) Annu Rev Cell Dev Biol 16, 653-99.

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 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.
SignalSilence is a registered trademark of Cell Signaling Technology, Inc.
SimpleChIP 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
#13987

c-Myc/N-Myc (D3N8F) Rabbit mAb

Western Blotting Image 1: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 c-Myc/N-Myc (D3N8F) 兔单克隆抗体,对不同细胞系提取物进行蛋白质印迹法分析。
Western Blotting Image 2: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 c-Myc/N-Myc (D3N8F) 兔单克隆抗体(上)或 β-Actin (D6A8) 兔单克隆抗体 #8457(下),对未处理 (-) 或用 JQ1(1 μM;72 小时;+)处理的 HT-29 细胞提取物,进行蛋白质印迹法分析。正如预期,BET 布罗莫结构域抑制剂 JQ1,可抑制 c-Myc 的表达。
Western Blotting Image 3: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 cMyc/N-Myc (D3N8F) 兔单克隆抗体(左)、N-Myc (D1V2A) 兔单克隆抗体 #84406(中)或 β-Actin (13E5) 兔单克隆抗体 #4970(右),对 K562 [c-Myc (+) / N-Myc (-)] 和 IMR-32 [c-Myc (-) / N-Myc (+)] 细胞系提取物进行蛋白质印迹法分析。
Western Blotting Image 4: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 cMyc/N-Myc (D3N8F) 兔单克隆抗体(上)或 β-Actin (D6A8) 兔单克隆抗体 #8457(下),对转染了 100 nM SignalSilence® 对照 siRNA(非偶联)#6568 (-) 或 SignalSilence® c-Myc siRNA I #6341 (+) 的 HeLa 细胞进行蛋白质印迹法分析。cMyc/N-Myc (D3N8F) 兔单克隆抗体确认了 c-Myc 表达的沉默,而 β-Actin (D6A8) 兔单克隆抗体则被用作上样对照。
Immunofluorescence Image 1: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 cMyc/N-Myc (D3N8F) 兔单克隆抗体(绿色),对未处理(左)或用布罗莫结构域抑制剂 JQ1(1 μM,72 小时;右)处理的 HT-29 细胞,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: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 cMyc/N-Myc (D3N8F) 兔单克隆抗体对未经(绿色)或已经 JQ1(1uM,72 小时,37℃)(蓝色)处理的 HT29 细胞进行流式细胞分析。Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) 作为二抗。
Chromatin Immunoprecipitation Image 1: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 SimpleChIP® 酶解染色质免疫沉淀试剂盒(磁珠法)#9003,对 Daudi 细胞的交联染色质和 cMyc/N-Myc (D3N8F) 兔单克隆抗体,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。图片中显示横跨 NPM1 的结合,而 NPM1 是知名的 c-Myc 靶标基因(请参阅含有 ChIP-qPCR 数据的附图)。
Chromatin Immunoprecipitation Image 2: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 SimpleChIP® 酶解染色质免疫沉淀试剂盒(磁珠法)#9003,对 Daudi 细胞的交联染色质和 cMyc/N-Myc (D3N8F) 兔单克隆抗体,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 5(上图)内的结合,包括 c-Myc 的已知靶标基因 NPM1(下图)(参见包含 ChIP-qPCR 数据的其他结果图)。
Chromatin Immunoprecipitation Image 3: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 SimpleChIP® 酶解染色质免疫沉淀试剂盒(磁珠法)#9003,对 Daudi 细胞的交联染色质和 cMyc/N-Myc (D3N8F) 兔单克隆抗体或正常兔 IgG #2729,进行染色质免疫沉淀分析。使用 human ATF4 promoter primers、SimpleChIP® Human NPM1 Intron 1 Primers #4779 和 SimpleChIP® Human α Satellite Repeat Primers #4486,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: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652,对 Daudi 细胞和 c-Myc/N-Myc (D3N8F) Rabbit mAb 进行 CUT&RUN 检测。‚使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。该图显示 c-MYC/N-Myc 的已知靶标基因 ABR 内的结合(参见包含 CUT&RUN-qPCR 数据的其他结果图)。
CUT and RUN Image 2: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652,对 Daudi 细胞和 c-Myc/N-Myc (D3N8F) Rabbit mAb 进行 CUT&RUN 检测。‚使用 DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。该图显示在染色体 1(上图)内的结合,包括 c-MYC/N-Myc 的已知靶基因 ABR(下图)(参见包含 CUT&RUN-qPCR 数据的其他结果图)。
CUT and RUN Image 3: c-Myc/N-Myc (D3N8F) Rabbit mAb Expand Image
使用 CUT&RUN Assay Kit #86652,对 Daudi 细胞和 c-Myc/N-Myc (D3N8F) Rabbit mAb 或 Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362 进行 CUT&RUN 检测。使用 human HNRNPL intron 1 primers、human ABR intron 1 primers 和 SimpleChIP® Human α Satellite Repeat Primers #4486,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。