Revision 3

#87182Store 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

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

18

Source/Isotype:

Rabbit IgG

UniProt ID:

#P25791

Entrez-Gene Id:

4005

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 10^6 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

Application Dilution
Western Blotting 1:1000
Immunofluorescence (Immunocytochemistry) 1:1600 - 1:6400
Flow Cytometry (Fixed/Permeabilized) 1:400 - 1:1600
Chromatin IP 1:100
Chromatin IP-seq 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.

Specificity / Sensitivity

LMO2 (E8K6I) Rabbit mAb recognizes endogenous levels of total LMO2 protein. This antibody also recognizes a non-specific band of unknown origin at 78 kDa. Testing of LMO2 (E8K6I) Rabbit mAb with positive and negative models by Flow Cytometry and Immunofluorescence corroborated specificity of this antibody.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human LMO2 protein.

Background

LIM-domain-only protein 2 (LMO2) is a transcriptional regulator that was first identified as a proto-oncogene activated by chromosomal translocations in T-cell acute lymphocytic leukemia (T-ALL) (1). Since then, LMO2 has been found to be a master regulator essential for erythroid development, as evidenced by homozygous LMO2 knockouts resulting in embryonic lethality in mice (2). LMO2 is an intrinsically unstructured protein that does not bind DNA directly, but rather acts as a scaffolding protein that recruits various transcription factors via its tandem cysteine-rich LIM domains to form a multi-protein DNA binding complex (3,4). The LMO2 complex plays a major role in hematopoiesis and was originally shown to consist of the transcription factors TAL1, E47, and GATA-1 in erythroid lineage cells, but variations of this complex may contain alternate transcription factors including LYL1, TAL2, GATA-2, and GATA-3 (5-9). The LMO2 complex also requires interaction with LIM-domain binding protein 1 (LDB1), which is necessary for LMO2 protein stability (10-11). In addition to hematopoietic tissue, LMO2 is also expressed in embryonic brain tissue, where it associates with BEX2 and the transcription factor NSCL-2 (12). Aberrant LMO2 expression is observed in several types of hematopoietic cancers, including large diffuse B-cell lymphoma (DLBCL), B-cell acute leukemia, (B-ALL), acute myeloid leukemia (AML), and T-ALL (13-16). LMO2-mediated T-ALL is primarily caused by the translocation t(11;14)(p13;q11) with the TCRD/A gene from chromosome 14q11, or the t(7;11)(q35;p13) translocation involving TCRB from 7q35 (1). In addition to hematopoietic cancers, overexpression of LMO2 in prostate stromal cells also facilitates prostate cancer progression by inducing expression of Interleukin-11 (IL-11), which stimulates STAT3 signaling in these cells (17).

  1. Boehm, T. et al. (1991) Proc Natl Acad Sci U S A 88, 4367-71.
  2. Warren, A.J. et al. (1994) Cell 78, 45-57.
  3. Chambers, J. and Rabbitts, T.H. (2015) Open Biol 5, 150062.
  4. Lécuyer, E. et al. (2007) J Biol Chem 282, 33649-58.
  5. Valge-Archer, V.E. et al. (1994) Proc Natl Acad Sci U S A 91, 8617-21.
  6. Wadman, I. et al. (1994) EMBO J 13, 4831-9.
  7. Wadman, I.A. et al. (1997) EMBO J 16, 3145-57.
  8. Osada, H. et al. (1995) Proc Natl Acad Sci U S A 92, 9585-9.
  9. Ono, Y. et al. (1998) Mol Cell Biol 18, 6939-50.
  10. Agulnick, A.D. et al. (1996) Nature 384, 270-2.
  11. Layer, J.H. et al. (2016) Mol Cell Biol 36, 488-506.
  12. Han, C. et al. (2005) Nucleic Acids Res 33, 6555-65.
  13. Natkunam, Y. et al. (2007) Blood 109, 1636-42.
  14. de Boer, J. et al. (2011) Leukemia 25, 321-30.
  15. Atay, M.H. et al. (2013) Histopathology 63, 293-4.
  16. Patel, J.L. et al. (2014) Histopathology 64, 226-33.
  17. Jiang, C.Y. et al. (2016) Oncotarget 7, 26247-58.

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

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 3
#87182

LMO2 (E8K6I) Rabbit mAb

Western Blotting Image 1: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 LMO2 (E8K6I) Rabbit mAb 对 TF-1、SET-2 和 K-562 细胞的提取物进行蛋白印迹分析。
Western Blotting Image 2: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 LMO2 (E8K6I) Rabbit mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图)对不同细胞系的提取物进行蛋白印迹分析。正如预期的一样,A549 和 Jurkat 细胞的 LMO2 表达呈阴性。
Immunofluorescence Image 1: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 LMO2 (E8K6I) Rabbit mAb(绿色)对 SET-2 细胞(左图,阳性)和 A549 细胞(右图,阴性)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。样本用 ProLong® Gold Antifade Reagent with DAPI #8961(蓝色)封片。
Flow Cytometry Image 1: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 LMO2 (E8K6I) Rabbit mAb(实线)或浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(虚线)对 Jurkat 细胞(蓝色,阴性)和 TF-1 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: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005 对 TF1 细胞中的交联染色质和 LMO2 (E8K6I) Rabbit mAb 进行染色质免疫沉淀分析。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。该图显示在 LYL1 基因内的结合。
Chromatin Immunoprecipitation Image 2: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005 对 TF1 细胞中的交联染色质和 LMO2 (E8K6I) Rabbit mAb 进行染色质免疫沉淀分析。使用 DNA Library Prep Kit for Illumina Systems (ChIP-seq, CUT&RUN) #56795 制备 DNA 库。该图显示在染色体 19(上图)内的结合,包括 LYL1 基因(下图)。
Chromatin Immunoprecipitation Image 3: LMO2 (E8K6I) Rabbit mAb Expand Image
使用 SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005 对 TF-1 细胞的交联染色质与 LMO2 (E8K6I) Rabbit mAb 或 Normal Rabbit IgG #2729 进行染色质免疫沉淀。使用 SimpleChIP® Human GYPA Promoter Primers #31805 和 SimpleChIP® Human α Satellite Repeat Primers #4486 进行实时 PCR 来对富集的 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。