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P ser824 kap 1

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The P-Ser824 KAP-1 is a lab equipment product designed for the detection and quantification of phosphorylated KAP-1 protein. It is a tool used in biological research applications.

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Lab products found in correlation

P h2ax ser139, by Merck Group (1 mentions) Pser345 chk1, by Cell Signaling Technology (1 mentions) Parp1, by Cell Signaling Technology (1 mentions) Odyssey infrared imaging system, by LI COR (1 mentions) β actin, by Merck Group (1 mentions) Alexa fluor 568, by Thermo Fisher Scientific (1 mentions) Anti chk2, by Cell Signaling Technology (1 mentions) Alexa fluor 680, by Thermo Fisher Scientific (1 mentions) Anti rabbit dylight 800, by Cell Signaling Technology (1 mentions) P p53 ser15, by Cell Signaling Technology (1 mentions) Anti α tubulin, by Cell Signaling Technology (1 mentions) Alexa fluor 488, by Thermo Fisher Scientific (1 mentions) Anti gapdh, by Merck Group (1 mentions) Alexa fluor 568 anti mouse igg, by Thermo Fisher Scientific (1 mentions) γh2ax, by Merck Group (1 mentions) Ubiqapture q, by Enzo Life Sciences (1 mentions) Gapdh, by Abcam (1 mentions)

4 protocols using p ser824 kap 1

1

Protein Extraction and Western Blotting for DNA Damage Signaling

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For total protein extract preparation, AMLMLL cells were collected upon treatment with ATRi (5 μM for 6 h) or HU (2 mM for 2 h), washed once with PBS and lysed in UREA buffer (8 M urea, 1 % Chaps, Tris-HCl 50 mM, pH 8.0) for 30 min at 4°C with agitation. Samples were resolved by SDS-PAGE and analysed by standard western blotting techniques. The following primary antibodies were used: p-Ser345 Chk1 (Cell Signaling), p-Ser4/Ser8 RPA32 (Bethyl), p-Ser139 H2AX (Millipore), H2AX (Abcam), p-SMC1 (Monoclonal Antibody Unit, CNIO), p-Ser824 KAP-1 (Bethyl) and PARP1 (Cell Signaling). Alexa Fluor 680- or 800- conjugated secondary antibodies (Life Technologies) were used for detection with a LI-COR Odyssey infrared imaging system (LI-COR Biosciences).
Morgado-Palacin I., Day A., Murga M., Lafarga V., Anton M.E., Tubbs A., Chen H.T., Ergan A., Anderson R., Bhandoola A., Pike K.G., Barlaam B., Cadogan E., Wang X., Pierce A.J., Hubbard C., Armstrong S.A., Nussenzweig A, & Fernandez-Capetillo O. (2016). Targeting the kinase activities of ATR and ATM exhibits therapeutic potential in a mouse model of MLL-rearranged AML. Science signaling, 9(445), ra91.
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2

Immunoblotting of DNA Damage Proteins

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Cells were lysed in cell extraction buffer (50 mM Tris pH 7.4, 150 mM NaCl, 2 mM EGTA, 2 mM EDTA, 25 mM NaF, 0.1 mM Na3VO4, 0.1 mM PMSF, 2 mg/mL Leupeptin, 2 mg/mL Aprotinin, 0.2% Triton X-100, 0.3% NonidetP-40) for 30 min on ice and centrifuged at 16100 rcf for 15 min. Protein extracts were separated through SDS-PAGE followed by immunoblotting. Primary antibodies against the following proteins were used: ATM (rabbit monoclonal, Epitomics, 1:1,000), DNA-PKcs (mouse monoclonal, Calbiochem, 1:500), pSer824-KAP1 (rabbit monoclonal, Bethyl Laboratories, 1:5,000), RNF168 (rabbit polyclonal, GeneTex, 1:1,000), and β-Actin (mouse monoclonal, Sigma, 1:3,000). Anti-mouse and anti-rabbit horseradish peroxidases labeled secondary antibodies were purchased from GE Healthcare. Enhanced chemiluminescence (Dura ECL, Thermo Scientific/Pierce) was used for visualization of immunoreactive bands.
Pietrucha B., Heropolitańska-Pliszka E., Geffers R., Enßen J., Wieland B., Bogdanova N.V, & Dörk T. (2017). Clinical and Biological Manifestation of RNF168 Deficiency in Two Polish Siblings. Frontiers in Immunology, 8, 1683.
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3

DNA Damage Response Protein Analysis

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Primary antibodies include p-ATM Ser1981 (Epitomics), ATM (Genetex), p-KAP-1 Ser824 (Bethyl), KAP-1 (CalBiochem), p-p53 Ser15 (Cell Signaling), anti-p53 (Calbiochem or Santa Cruz DO-1), anti-p(T68)-CHK2 (Cell Signaling), anti-CHK2 (Cell Signaling), cleaved caspase 3 rabbit mAb (5A1E), anti-α-tubulin (Cell Signaling), and anti-GAPDH (EMD Millipore). Secondary antibodies for westerns were anti-rabbit Dylight-800 (Cell Signaling) and anti-mouse Alexa Fluor-680 (Invitrogen). Secondary antibodies for immunofluorescence were AlexaFluor-488, AlexaFluor-568, and AlexaFluor-647 anti-IgG (Invitrogen).
Karlin J., Allen J., Ahmad S.F., Hughes G., Sheridan V., Odedra R., Farrington P., Cadogan E.B., Riches L.C., Garcia-Trinidad A., Thomason A.G., Patel B., Vincent J., Lau A., Pike K.G., Hunt T.A., Sule A., Valerie N.C., Biddlestone-Thorpe L., Kahn J., Beckta J.M., Mukhopadhyay N., Barlaam B., Degorce S.L., Kettle J., Colclough N., Wilson J., Smith A., Barrett I.P., Zheng L., Zhang T., Wang Y., Chen K., Pass M., Durant S.T, & Valerie K. (2018). Orally bioavailable and blood-brain-barrier penetrating ATM inhibitor (AZ32) radiosensitizes intracranial gliomas in mice. Molecular cancer therapeutics, 17(8), 1637-1647.
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4

Antibody-based Protein Modification Analysis

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The following primary antibodies were used: TRIM33 (#90,051, CST, Leiden, The Netherlands), pCHK1Ser345 (#2348, CST), CHK1 (PA5-96,749, Invitrogen), pKAP1Ser824 (A300-767A, Bethyl Laboratories), KAP1 (A700-014, Bethyl Laboratories, Cambridge, UK), γH2AX (SAB5600038, Sigma Aldrich, Dorset, UK), GAPDH (ab8245, Abcam, Cambridge, UK), ALC1 (av51324, Abcam), K48-linkage specific polyubiquitination (#8081, CST) and 53BP1 (sc-517281, Santa Cruz Biotechnology, Texas, USA). The following secondary antibodies were used for Western blotting, Anti-Mouse IgG HRP-linked (#7076P2, CST) and Anti-Rabbit IgG HRP-linked (#7074P2, CST), and Anti-Mouse IgG Alexa Fluor 568 (A-11004, ThermoFisher Scientific) was used for immunofluorescence staining. Ubiquitinated proteins were isolated using UbiQapture-Q (Enzo Life Sciences, Exeter, UK) to bind both mono- and poly-ubiquitinated proteins independent of lysine chain linkage.
McAvera R.M., Morgan J.J., Herrero A.B., Mills K.I, & Crawford L.J. (2024). TRIM33 loss in multiple myeloma is associated with genomic instability and sensitivity to PARP inhibitors. Scientific Reports, 14, 8797.
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