Revision 3

#3287Store 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, IP, IHC-P, IF-IC, FC-FP

REACTIVITY:

H

SENSITIVITY:

Endogenous

MW (kDa):

258, 110, 50-80

Source/Isotype:

Rabbit IgG

UniProt ID:

#P08922

Entrez-Gene Id:

6098

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10 - 1:50
Immunoprecipitation 1:50
Immunohistochemistry (Paraffin) 1:250
Immunofluorescence (Immunocytochemistry) 1:200 - 1:800
Flow Cytometry (Fixed/Permeabilized) 1:200 - 1:800

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

ROS1 (D4D6®) Rabbit mAb recognizes endogenous levels of total ROS1 protein. This antibody does not cross-react with other related proteins when analyzed by western blot. Please note that staining may be observed in ROS1 rearranged lung carcinomas, macrophages/giant cells, reactive type II pneumocyte hyperplasia, and the epithelium in areas of bronchiolar metaplasia. Staining of unknown specificity has been observed in cholangiocarcinoma, hepatocellular carcinoma, and kidney tissues.

Species Reactivity:

Human

Source / Purification

Monoclonal antibody is produced by immunizing animals with a protein corresponding to residues in the carboxy terminal domain of the human ROS1 protein.

Background

ROS1, an orphan receptor tyrosine kinase of the insulin receptor family, was initially identified as a homolog of v-ros from the UR2 sarcoma virus (1). ROS1 consists of a large extracellular domain that is composed of six fibronectin repeats, a transmembrane domain, and a C-terminal kinase domain. Being an orphan receptor, the functions of ROS1 are not well known, though it has been shown to play an important role in differentiation of epididymal epithelium (2). The first oncogenic fusion of ROS1, FIG-ROS1, was initially identified by research studies in glioblastoma (3), and subsequent studies have found this fusion in cholangiocarcinoma (4), ovarian cancer (5), and non-small cell lung cancer (NSCLC) (6). Investigators have found additional oncogenic ROS1 fusion proteins in NSCLC (at a frequency of ~1.6%), where the ROS1 kinase domain is fused to the amino-terminal region of several different proteins, including CD74 and SLC34A2 (6-8). ROS1 fusion proteins activate the SHP-2 phosphatase, PI3K/Akt/mTOR, Erk, and Stat3 pathways (3,4,9). There are two autophosphorylation sites (Tyr2274, Tyr2334) downstream of the kinase domain of ROS1, either of which may serve as biomarkers of ROS1 kinase activity, including that of ROS1 fusion proteins (10).

  1. Matsushime, H. et al. (1986) Mol Cell Biol 6, 3000-4.
  2. Yeung, C.H. et al. (1999) Biol Reprod 61, 1062-9.
  3. Charest, A. et al. (2003) Genes Chromosomes Cancer 37, 58-71.
  4. Gu, T.L. et al. (2011) PLoS One 6, e15640.
  5. Birch, A.H. et al. (2011) PLoS One 6, e28250.
  6. Rimkunas, V.M. et al. (2012) Clin Cancer Res 18, 4449-57.
  7. Rikova, K. et al. (2007) Cell 131, 1190-203.
  8. Stumpfova, M. and Jänne, P.A. (2012) Clin Cancer Res 18, 4222-4.
  9. Jun, H.J. et al. (2012) Cancer Res 72, 3764-74.
  10. Zou, H.Y. et al. (2015) Proc Natl Acad Sci U S A 112, 3493-8.

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 W-S: Simple Western™ IP: Immunoprecipitation IHC-P: Immunohistochemistry (Paraffin) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

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

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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
#3287

ROS1 (D4D6®) Rabbit mAb

Western Blotting Image 1: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 ROS1 (D4D6®) Rabbit mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对 HCC78 (SLC34A2-ROS1)、U-118 MG (FIG-ROS1) 和 HeLa(ROS1 阴性)细胞的提取物进行蛋白质印迹分析。注:HCC78 细胞表达 85、70 和 59 kDa 的 SLC34A2-ROS1 融合蛋白 (7)。
Western Blotting Image 2: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 ROS1 (D4D6®) Rabbit mAb(上图)或 β-Actin (D6A8) Rabbit mAb #8457(下图),对 转染空载 (-) 或转染野生型 ROS1 (hROS1; +) 表达载体的 293T 细胞的提取物进行蛋白质印迹分析。
Western Blotting Image 1: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 ROS1 (D4D6® ) Rabbit mAb #3287 对 HCC-78 细胞的裂解物 (1.0 mg/mL) 进行 Simple Western™ 分析。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。在还原条件下,使用 66-440 kDa 分离模块在 ProteinSimple(BioTechne 品牌)的 Jess™ Simple Western 仪器上进行该实验。
Immunoprecipitation Image 1: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(泳道 2)或 ROS1 (D4D6®) Rabbit mAb(泳道 3),对 HCC78 细胞提取物 ROS1 进行免疫沉淀。泳道 1 是 10% 输入对照。使用 ROS1 (D4D6®) Rabbit mAb 后再用 Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb #3678 和 Anti-mouse IgG, HRP-linked Antibody #7076 进行蛋白质印迹分析。
Immunohistochemistry Image 1: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 ROS1 (D4D6®) Rabbit mAb 对石蜡包埋的且显示不同定位的人肺癌细胞进行免疫组织化学分析。
No image available
Flow Cytometry Image 1: ROS1 (D4D6®) Rabbit mAb Expand Image
使用 ROS1 (D4D6®) Rabbit mAb(实线)或浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(虚线)对 HeLa 细胞(蓝色,阴性)和 HCC78 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 用作二抗。