Revision 1
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:

IF-IC, FC-FP

REACTIVITY:

H M R Mk

SENSITIVITY:

Endogenous

MW (kDa):

Source/Isotype:

Rabbit IgG

UniProt ID:

#Q96QV6

Entrez-Gene Id:

221613

Product Information

Product Usage Information

Application Dilution
Immunofluorescence (Immunocytochemistry) 1:200
Flow Cytometry (Fixed/Permeabilized) 1:50

Storage

Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.

Specificity / Sensitivity

Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) recognizes endogenous levels of histone H2A protein only when ubiquitinated at Lys119. The antibody does not cross-react with other ubiquitinated proteins or free ubiquitin.

Species Reactivity:

Human, Mouse, Rat, Monkey

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human histone H2A protein in which Lys119 is mono-ubiquitinated.

Product Description

This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 488 fluorescent dye and tested in-house for direct flow cytometric and immunofluorescent analysis in human and mouse cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb #8240.

Background

The nucleosome, made up of four core histone proteins (H2A, H2B, H3, and H4), is the primary building block of chromatin. Originally thought to function as a static scaffold for DNA packaging, histones have now been shown to be dynamic proteins, undergoing multiple types of posttranslational modifications, including acetylation, phosphorylation, methylation, and ubiquitination (1). Ubiquitin is a conserved 76 amino acid peptide unit that can be covalently linked to many cellular proteins by the ubiquitination process. Three components are involved in this protein-ubiquitin conjugation process. Ubiquitin is first activated by forming a thioester complex with the activation component E1; the activated ubiquitin is subsequently transferred to the ubiquitin-carrier protein E2, then from E2 to ubiquitin ligase E3 for final delivery to the epsilon-NH2 of the target protein lysine residue (2). Histone H2A is mono-ubiquitinated at Lys119 by the Polycomb Repressor Complex 1 (PRC1) and is critical for transcriptional silencing of the developmental HOX genes and X chromosome inactivation (3-6). PRC1 is composed of Bmi1 and RING1A (also RING1 or RNF1), both of which act to enhance the E3 ubiquitin ligase activity of the catalytic subunit RING1B (also RING2 or RNF2) (3,4). Histone H2A is also mono-ubiquitinated at Lys119 at sites of DNA damage. This mono-ubiquitination event requires the PRC1 components Bmi1 and RING1B, in addition to another E3 ubiquitin ligase RNF8, and contributes to subsequent recruitment of the BRCA1 complex, via binding of RAP80/UIMC1 (ubiquitin interactive motif containing 1 protein) (7-10).

  1. Peterson, C.L. and Laniel, M.A. (2004) Curr Biol 14, R546-51.
  2. Liu, F. and Walters, K.J. (2010) Trends Biochem Sci 35, 352-60.
  3. Wang, H. et al. (2004) Nature 431, 873-8.
  4. Cao, R. et al. (2005) Mol Cell 20, 845-54.
  5. de Napoles, M. et al. (2004) Dev Cell 7, 663-76.
  6. Fang, J. et al. (2004) J Biol Chem 279, 52812-5.
  7. Ginjala, V. et al. (2011) Mol Cell Biol 31, 1972-82.
  8. Bergink, S. et al. (2006) Genes Dev 20, 1343-52.
  9. Marteijn, J.A. et al. (2009) J Cell Biol 186, 835-47.
  10. Wu, J. et al. (2009) Mol Cell Biol 29, 849-60.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Applications Key

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.
XP is a registered trademark of Cell Signaling Technology, Inc.
This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is conditioned on the buyer using the purchased product solely in research conducted by the buyer, excluding contract research or any fee for service research, and the buyer must not (1) use this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; or (c) manufacturing or quality assurance or quality control, and/or (2) sell or transfer this product or its components for resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or [email protected].
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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Revision 1
#26498

Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)

Immunofluorescence Image 1: Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) Expand Image
使用 Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)(绿色)和 β-Actin (8H10D10) Mouse mAb #3700(红色),对 HeLa 细胞(左图)和 NIH/3T3 细胞(右图)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 2: Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) Expand Image
使用 Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)(绿色)对 HeLa 细胞进行免疫荧光分析。在免疫染色之前,用无肽(左图)、对应于泛素化的组蛋白H2A(K119)的肽(中图)或不对应非泛素化的组蛋白H2A(K119)的肽(右图)激发抗体。样本用 ProLong® Gold Antifade Reagent with DAPI #8961(蓝色)封片。
Immunofluorescence Image 3: Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) Expand Image
使用 Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) 对 HeLa 细胞进行免疫荧光分析。在免疫染色之前,用无肽(绿色)、与泛素化组蛋白 H2A(K119)(蓝色)对应的肽或与非泛素化组蛋白 H2A(K119)(红色)相对应的肽激发抗体(x 轴)浓度的增加。然后测量荧光强度以确定最佳工作浓度。用非泛素化肽确认靶标特异性观察到轻微阻断。
Flow Cytometry Image 1: Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) Expand Image
使用 Ubiquityl-Histone H2A (Lys119) (D27C4) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)(实线)或浓度匹配的 Rabbit (DA1E) mAb IgG XP® Isotype Control (Alexa Fluor® 488 Conjugate) #2975(虚线)对 HeLa 细胞(蓝色)或 C2C12 细胞(绿色)进行流式细胞分析。