RPA2 [p Ser4, p Ser8] Antibody

Western Blot: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Detection of Human Phospho RPA32 (S4/S8) by western blot. Samples: Whole cell lysate (50 ug) from HeLa cells treated with 100 uM EPE (+) or mock treated (-). Antibodies: Affinity purified rabbit anti-Phospho RPA32 (S4/S8) antibody NBP1-23017 used at 0.1 ug/ml. Lower panel: Rabbit anti-RPA32 antibody NB100-332. Detection: Chemiluminescence with an exposure time of 75 seconds.

Immunocytochemistry/Immunofluorescence: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Formaldehyde-fixed asynchronous HeLa cells. Untreated cells (left) and EPE treated cells (right). Antibody: Affinity purified rabbit anti- Phospho RPA32 (S4/S8) used at a dilution of 1:1,000 (1ug/ml). Detection: Red-fluorescent goat anti-rabbit IgG-heavy and light chain cross-adsorbed Antibody DyLight® 594 Conjugated used at a dilution of 1:100. Counterstain: Alexa Fluor (R) 488 phalloidin (green).

Immunohistochemistry-Paraffin: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Sample: FFPE section of human lung cancer. Antibody: Affinity purified rabbit anti-Phospho RPA32 (S4/S8) used at a dilution of 1:1,000 (1ug/ml). Detection: DAB

Western Blot: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Samples: Whole cell lysate (50 ug for WB in A; 1 mg for IP, 20% of IP loaded in B) from HeLa Cells that had been treated with EPE (+) or mock treated (-). In B, immunoprecipitated RPA32 was either treated with phosphatase (+) or mock treated (-). Antibodies: Phosphorylated RPA32 was detected using rabbit anti-phospho RPA32 Ser4/Ser8 antibody NBP1-23017 lot 2 at 1 ug/ml. Total RPA32 was detected using rabbit anti-RPA32 antibody NB100-332 at 1 ug/ml. RPA32 was immunoprecipitated using NB100-332 at 3 ug/mg lysate. Detection: Chemiluminescence with exposure times of 30 seconds for phospho RPA32 Ser4/Ser8 (A and B) and 1 second for total RPA32 (A and B).

Western Blot: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Detection of Human Phospho RPA32 (S4/S8) by Western Blot. Samples: Whole cell lysate (50 ug) from HeLa cells treated with 100 uM EPE for 4 hours (+) or mock treated (-) cells. Antibodies: Affinity purified rabbit anti-Phospho RPA32 (S4/S8) antibody NBP1-23017 used for WB at 0.1 ug/ml. Detection: Chemiluminescence with an exposure time of 30 seconds. Lower panel shows Westen Blot for total RPA32 using affinity purified rabbit anti-RPA32 antibody A300-244A at 0.1 ug/ml with an exposure time of 30 seconds.

Immunohistochemistry-Paraffin: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Sample: FFPE section of mouse colon carcinoma CT26. Antibody: Affinity purified rabbit anti-Phospho RPA32 (S4/S8) used at a dilution of 1:1,000 (1ug/ml). Detection: DAB

Immunoprecipitation: RPA2 [p Ser4, p Ser8] Antibody [NBP1-23017] - Detection of human Phospho RPA32 (S4/S8) by western blot of immunoprecipitates. Samples: Whole cell lysate (1.0 mg per IP reaction; 20% of IP loaded) from HeLa cells treated with EPE (+) and/or Lambda PP (+) or mock treated (-). Antibodies: RPA32 (S4/S8) was immunoprecipitated using affinity purified rabbit anti-RPA32 antibody NB100-332. For blotting immunoprecipitated Phospho RPA32 (S4/S8), rabbit anti-Phospho RPA32 (S4/S8) antibody NBP1-23017 was used at 0.1 ug/ml. Lower Panel: Rabbit anti-RPA antibody NB100-332. Detection: Chemiluminescence with an exposure time of 75 seconds.

• Reactivity: Hu, Mu, Pm-Or
• Applications: WB, Simple Western, ICC/IF, IHC, IHC-P, IP, KD
• Clonality: Polyclonal
• Host: Rabbit
• Conjugate: Unconjugated
• Concentration: 1.0 mg/ml

RPA2 [p Ser4, p Ser8] Antibody Summary

• Immunogen: Immunogen was a dually phosphorylated synthetic peptide, which represented a portion of human replication protein A2, 32 kDa surrounding phosphorylated serines that corresponded to positions 4 and 8 using the numbering given in entry NP_002937.1 (GeneID
• Modification: p Ser4, p Ser8
• Predicted Species: Orangutan (100%). Backed by our 100% Guarantee.
• Isotype: IgG
• Clonality: Polyclonal
• Host: Rabbit
• Gene: RPA2
• Purity: Immunogen affinity purified

Applications/Dilutions

• Dilutions:
  • Immunocytochemistry/Immunofluorescence 1:500 - 1:5000
  • Immunohistochemistry 1:500 - 1:2000
  • Immunohistochemistry-Paraffin 1:500 - 1:2000
  • Immunoprecipitation 2-10 ug/mg of lysate
  • Knockdown Validated
  • Simple Western
  • Western Blot 1:2000-1:10000
• Application Notes: Formaldehyde fixation is recommended. Permeabilization with Triton-X 100 is recommended for formaldehydefixed cells. Epitope retrieval with Tris-EDTA pH9.0 is recommended for FFPE tissue sections.
  Simple Western reported by an internal validation. Separated by Charge, antibody dilution of 1:50.

Packaging, Storage & Formulations

• Storage: Store at 4C. Do not freeze.
• Buffer: Tris-Citrate/Phosphate (pH 7.0 - 8.0)
• Preservative: 0.09% Sodium Azide
• Concentration: 1.0 mg/ml
• Purity: Immunogen affinity purified

Alternate Names for RPA2 [p Ser4, p Ser8] Antibody

• REPA2
• Replication factor A protein 2
• replication protein A 32 kDa subunit
• Replication protein A 34 kDa subunit
• replication protein A2 (32kD)
• replication protein A2, 32kDa
• RF-A protein 2
• RP-A p32
• RP-A p34
• RPA2
• RPA32
• RPA34

Background

Replication protein A (RPA) is a multisubunit complex that carries out DNA mismatch repair (MMR) in association with MSH2 (a subunit of human MutS heterodimers), MLH1 (a subunit of human MutL heterodimers), PCNA, and DNA polymerase-delta. RPA is composed of a heterotrimer that includes subunits of 70 kDa (RPA1), 32kDa (RPA2), and 14kDa (RPA3). RPA2, the 32kDa subunit, is phosphorylated by the cdc2 family of kinases when cells enter S-phase and in response to DNA damage by ATM, ATR, and DNA-PK.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for RPA2 Antibody (NBP1-23017)(37)

Publications using NBP1-23017

• de Krijger I, FOhr B, PErez SH et al. MAD2L2 dimerization and TRIP13 control shieldin activity in DNA repair Nature communications Sep 14 2021 [PMID: 34521823] (WB, Human)
• Kumar S, Buon L, Talluri S et al. Integrated genomics and comprehensive validation reveal drivers of genomic evolution in esophageal adenocarcinoma Communications biology May 24 2021 [PMID: 34031527]
• Li H, Ma L, Bian X et al. FK228 sensitizes radioresistant small cell lung cancer cells to radiation Clinical epigenetics Feb 25 2021 [PMID: 33632300] (Human)
• Bian X, Wang X, Zhang Q et al. The MYC Paralog-PARP1 Axis as a Potential Therapeutic Target in MYC Paralog-Activated Small Cell Lung Cancer Front. Oncol. Oct 8 2020 [PMID: 33134168] (WB, Human)
• Bartek J, Merchut-Maya JM, Maya-Mendoza A et al. Cancer cell stemness, responses to experimental genotoxic treatments, cytomegalovirus protein expression and DNA replication stress in pediatric medulloblastomas Cell Cycle Feb 13 2020 [PMID: 32054408] (Human)
• Wang H, Zhang S et al. Synergistic lethality between PARP-trapping and alantolactone-induced oxidative DNA damage in homologous recombination-proficient cancer cells. Oncogene Jan 4 2020 [PMID: 32029902] (ICC/IF, Human)
• Fagan-Solis KD, Simpson DA, Kumar RJ et al. A P53-Independent DNA Damage Response Suppresses Oncogenic Proliferation and Genome Instability Cell Rep Feb 4 2020 [PMID: 32023457] (ICC/IF, Mouse)
• Warren N, Eastman A. Inhibition of checkpoint kinase 1 following gemcitabine-mediated S phase arrest results in CDC7- and CDK2-dependent replication catastrophe. J. Biol. Chem. Dec 20 2018 [PMID: 30573684] (WB, Human)
• Simonetta M, de Krijger I, Serrat J et al. H4K20me2 distinguishes pre-replicative from post-replicative chromatin to appropriately direct DNA repair pathway choice by 53BP1-RIF1-MAD2L2. Cell Cycle. [PMID: 29160738] (WB, Human)
• Qu J, Sun W, Zhong J et al. Correction: Phosphoglycerate mutase 1 regulates dNTP pool and promotes homologous recombination repair in cancer cells J. Cell Biol. 2017 Jul 24 [PMID: 28739677] (WB, Human)
• Qu J, Sun W, Zhong J et al. Phosphoglycerate mutase 1 regulates dNTP pool and promotes homologous recombination repair in cancer cells J Cell Biol. 2016 Jul 03 [PMID: 28122957] (WB, ICC/IF, KD, Human)
• Sakurikar N, Thompson R, Montano R, Eastman A A subset of cancer cell lines is acutely sensitive to the Chk1 inhibitor MK-8776 as monotherapy due to CDK2 activation in S phase. Oncotarget 2016 Jan 12 [PMID: 26595527] (WB)
• Fagan-Solis K, Simpson D, Kumar R et al. A P53-Independent DNA Damage Response Suppresses Oncogenic Proliferation and Genome Instability SSRN Journal Jul 3 2019
• Thompson R, Montano R, Eastman A et al. The Mre11 nuclease is critical for the sensitivity of cells to Chk1 inhibition. PLoS One 2012 [PMID: 22937147]
• Kousholt AN, Fugger K, Hoffmann S et al. CtIP-dependent DNA resection is required for DNA damage checkpoint maintenance but not initiation. J Cell Biol 2012 Jun [PMID: 22733999]
• Punchihewa C, Inoue A, Hishiki A et al. Identification of small molecule proliferating cell nuclear antigen (PCNA) inhibitor that disrupts interactions with PIP-box proteins and inhibits DNA replication. J Biol Chem 2012 Apr [PMID: 22383522]
• Regairaz M, Zhang Y-W, Fu H et al. Mus81-mediated DNA cleavage resolves replication forks stalled by topoisomerase I-DNA complexes. J Cell Biol 28195(5):739-749. 2011 Nov. [PMID: 22123861]
• Xu LH, Huang M, Fang SG, Liu DX. Coronavirus infection induces DNA replication stress partly through interaction of its nonstructural protein 13 with the p125 subunit of DNA polymerase delta. Journal of Biological Chemistry 286 (45):39546-39559. 2011 [PMID: 21918226]
• Liu S, Chu J, Yucer N et al. RING finger and WD repeat domain 3 (RFWD3) associates with replication protein A (RPA) and facilitates RPA-mediated DNA damage response. J.Biol.Chem 286 (25):22314-22322. 2011 [PMID: 21558276]
• Baldeyron C, Soria G, Roche D et al. HP1alpha recruitment to DNA damage by p150CAF-1 promotes homologous recombination repair. J Cell Biol 2011 Apr [PMID: 21464229]
• Du G, Dutta N, Lashmit P et al. Alternative splicing of the human cytomegalovirus major immediate-early genes affects infectious-virus replication and control of cellular cyclin-dependent kinase. J Virol 2011 Jan [PMID: 21068259]
• Chen J, Luo Q, Yuan Y et al. Pygo2 associates with MLL2 histone methyltransferase and GCN5 histone acetyltransferase complexes to augment Wnt target gene expression and breast cancer stem-like cell expansion. Mol.Cell.Biol. 30 (24):5621-5635. 2010 [PMID: 20937768]
• Shi W, Feng Z, Zhang J et al. The role of RPA2 phosphorylation in homologous recombination in response to replication arrest. Carcinogenesis 2010 Jun [PMID: 20130019]
• Glauser DL, Seyffert M, Strasser R et al. Inhibition of herpes simplex virus type 1 replication by adeno-associated virus rep proteins depends on their combined DNA-binding and ATPase/helicase activities. J Virol 2010 Apr [PMID: 20106923]
• Cerqueira A, Santamaria D, Martinez-Pastor B et al. Overall Cdk activity modulates the DNA damage response in mammalian cells. J Cell Biol 2009 Dec [PMID: 19995934]
• Stephan H, Concannon C, Kremmer E et al. Ionizing radiation-dependent and independent phosphorylation of the 32-kDa subunit of replication protein A during mitosis. Nucleic Acids Res 2009 Oct [PMID: 19671522]
• Huertas P, Jackson SP. Human CtIP mediates cell cycle control of DNA end resection and double strand break repair. J Biol Chem 2009 Apr [PMID: 19202191]
• Carson CT, Orazio NI, Lee DV et al. Mislocalization of the MRN complex prevents ATR signaling during adenovirus infection. EMBO J 2009 Mar [PMID: 19197236]
• Gravel S, Chapman JR, Magill C et al. DNA helicases Sgs1 and BLM promote DNA double-strand break resection. Genes Dev 2008 Oct [PMID: 18923075]
• Kranz D, Dohmesen C, Dobbelstein M et al. BRCA1 and Tip60 determine the cellular response to ultraviolet irradiation through distinct pathways. J Cell Biol 2008 Jul [PMID: 18625847]
• Fragkos M, Breuleux M, Clement N et al. Recombinant adeno-associated viral vectors are deficient in provoking a DNA damage response. J Virol 2008 Aug [PMID: 18463154]
• Manthey KC, Opiyo S, Glanzer JG et al. NBS1 mediates ATR-dependent RPA hyperphosphorylation following replication-fork stall and collapse. J Cell Sci 2007 Dec [PMID: 18003706]
• Morency E, Sabra M, Catez F et al. A novel cell response triggered by interphase centromere structural instability. J Cell Biol 2007 Jun [PMID: 17548509]
• Olson E, Nievera CJ, Klimovich V et al. RPA2 is a direct downstream target for ATR to regulate the S-phase checkpoint. J Biol Chem 2006 Dec [PMID: 17035231]
• Cruet-Hennequart S, Coyne S, Glynn MT et al. UV-induced RPA phosphorylation is increased in the absence of DNA polymerase eta and requires DNA-PK. DNA Repair (Amst) 2006 Apr [PMID: 16520097]
• Wu X, Shell SM, Zou Y et al. Interaction and colocalization of Rad9/Rad1/Hus1 checkpoint complex with replication protein A in human cells. Oncogene 2005 Jul [PMID: 15897895]
• Vassin VM, Wold MS, Borowiec JA et al. Replication protein A (RPA) phosphorylation prevents RPA association with replication centers. Mol Cell Biol 2004 Mar [PMID: 14966274]

Bioinformatics

• Gene Symbol: RPA2
• Entrez:
  • 6118(Human)
• Uniprot:
  • NP_002937.1(Human)