Revision 8
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-F, IF-IC, FC-FP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

110

Source/Isotype:

Rabbit IgG

UniProt ID:

#P06400

Entrez-Gene Id:

5925

Product Information

Product Usage Information

Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10
Immunoprecipitation 1:200
Immunohistochemistry (Paraffin) 1:200 - 1:800
Immunofluorescence (Frozen) 1:400 - 1:800
Immunofluorescence (Immunocytochemistry) 1:800 - 1:3200
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.

For a carrier-free (BSA and azide free) version of this product see product #41359.

Specificity / Sensitivity

Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb recognizes endogenous levels of Rb protein only when phosphorylated at Ser807, Ser811, or at both sites. This antibody does not cross-react with Rb phosphorylated at Ser608.

Species Reactivity:

Human, Mouse, Rat, Monkey

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ser807/811 of human Rb protein.

Background

The retinoblastoma tumor suppressor protein Rb regulates cell proliferation by controlling progression through the restriction point within the G1-phase of the cell cycle (1). Rb has three functionally distinct binding domains and interacts with critical regulatory proteins including the E2F family of transcription factors, c-Abl tyrosine kinase, and proteins with a conserved LXCXE motif (2-4). Cell cycle-dependent phosphorylation by a CDK inhibits Rb target binding and allows cell cycle progression (5). Rb inactivation and subsequent cell cycle progression likely requires an initial phosphorylation by cyclin D-CDK4/6 followed by cyclin E-CDK2 phosphorylation (6). Specificity of different CDK/cyclin complexes has been observed in vitro (6-8) and cyclin D1 is required for Ser780 phosphorylation in vivo (9).

  1. Sherr, C.J. (1996) Science 274, 1672-7.
  2. Nevins, J.R. (1992) Science 258, 424-9.
  3. Welch, P.J. and Wang, J.Y. (1993) Cell 75, 779-90.
  4. Hu, Q.J. et al. (1990) EMBO J 9, 1147-55.
  5. Knudsen, E.S. and Wang, J.Y. (1997) Mol Cell Biol 17, 5771-83.
  6. Lundberg, A.S. and Weinberg, R.A. (1998) Mol Cell Biol 18, 753-61.
  7. Connell-Crowley, L. et al. (1997) Mol Biol Cell 8, 287-301.
  8. Kitagawa, M. et al. (1996) EMBO J 15, 7060-9.
  9. Geng, Y. et al. (2001) Proc Natl Acad Sci USA 98, 194-9.

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-F: Immunofluorescence (Frozen) 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

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 8
#8516

Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb

Western Blotting Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb(上)或 Rb (4H1) Mouse mAb #9309(下),对未经处理 (-) 或经小牛小肠磷酸酶 (CIP) 和 λ 磷酸酶 (+) 处理的 MCF7 细胞提取物进行蛋白质印迹分析。
Western Blotting Image 2: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb,对 WI-38 细胞提取物进行蛋白质印迹分析,而 WI-38 细胞经血清饥饿 3 天 (-) 或血清饥饿 3 天后用 10% 的血清处理 2 天 (+)。
Western Blotting Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
Simple Western™ analysis of lysates (0.1 mg/mL) from MCF-7 cells using Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb #8516. 虚拟泳道视图(左图)显示一抗稀释比例为 1:10 时的靶标条带(如图所示)。The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 dilution of primary antibody. 在还原条件下,使用 12-230 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: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Rabbit (DA1E) mAb IgG XP® Isotype Control #3900(泳道 2)或 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb(泳道 3),对 Cos 细胞提取物磷酸 Rb (Ser807/811) 进行免疫沉淀。泳道 1 是 10% 输入对照。使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb 进行蛋白质印迹分析。
Immunohistochemistry Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb 对石蜡包埋的人结肠癌进行免疫组织化学分析。
Immunohistochemistry Image 2: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb,对对照性(左)或经 λ 磷酸酶处理(右)的石蜡包埋的肺癌进行免疫组织化学分析。
Immunohistochemistry Image 3: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb 对石蜡包埋的小鼠脾脏细胞进行免疫组织化学分析。
Immunohistochemistry Image 4: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
在对照肽(左)或抗原特异性肽(右)存在的情况下,使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb,对石蜡包埋的人卵巢浆液性腺癌进行免疫组织化学分析。
Immunofluorescence Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse small intestine labeled with Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb (left, green) and co-labeled with DyLight 554 Phalloidin #13054 (right, red) and ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
Immunofluorescence Image 2: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse testis labeled with Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb (left, green) and co-labeled with DyLight 554 Phalloidin #13054 (right, red) and ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
Immunofluorescence Image 3: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of fixed frozen mouse thymus labeled with Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb (left, green) and co-labeled with DyLight 554 Phalloidin #13054 (right, red) and ProLong® Gold Antifade Reagent with DAPI #8961 (right, blue).
Immunofluorescence Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/Ser811) (D20B12) XP® Rabbit mAb(绿色),对未经处理(上)或经 λ 磷酸酶处理(下)的 MCF7(左)和 BT-549(右)细胞进行共聚焦免疫荧光分析。使用 DY-554 phalloidin(红色)标记肌动蛋白纤丝。蓝色伪彩 = DRAQ5® #4084(DNA 荧光染料)。
Flow Cytometry Image 1: Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb Expand Image
使用 Phospho-Rb (Ser807/811) (D20B12) XP® Rabbit mAb 和 Propidium Iodide (PI/RNase Staining Solution #4087对 Jurkat 细胞进行流式细胞分析,以测量 DNA 含量。Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 用作二抗。