Anti pchk2 t68

Manufactured by Cell Signaling Technology

Anti-pChk2 (T68) is a primary antibody that recognizes the phosphorylated form of Chk2 at threonine 68. Chk2 is a serine/threonine protein kinase that is activated in response to DNA damage.

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Lab products found in correlation

Anti chk2, by Cell Signaling Technology (2 mentions) Anti pchk1 s345, by Cell Signaling Technology (2 mentions) Bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Anti egfr, by Santa Cruz Biotechnology (1 mentions) Protease inhibitor, by Thermo Fisher Scientific (1 mentions) Anti 53bp1, by Cell Signaling Technology (1 mentions) Anti p akt, by Cell Signaling Technology (1 mentions) Anti p erk t202 y204, by Cell Signaling Technology (1 mentions) Anti met c 12, by Santa Cruz Biotechnology (1 mentions) Ecl western blotting kit, by Merck Group (1 mentions) Anti pdgfrβ, by Cell Signaling Technology (1 mentions) Pvdf membrane, by Merck Group (1 mentions) Anti gapdh, by Merck Group (1 mentions) Dynabead, by Thermo Fisher Scientific (1 mentions) Benzonase, by Merck Group (1 mentions) Anti chk1, by Cell Signaling Technology (1 mentions) Anti β actin, by Thermo Fisher Scientific (1 mentions) Anti vinculin, by Abcam (1 mentions) Anti 53bp1, by Thermo Fisher Scientific (1 mentions) Anti chk1, by Santa Cruz Biotechnology (1 mentions)

Close spelling variants of this product

Rabbit anti-pChk2 (T68) PCHK2 T68

4 protocols using anti pchk2 t68

Western Blot Analysis of Signaling Proteins

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Cells were lysed in the RIPA buffer (P89901, Thermo Scientific) supplemented with protease inhibitors (P1862209, Thermo Scientific). The protein concentration was detected using a BCA protein assay kit (Thermo Scientific). For western blot, 40 μg protein was separated by 8% SDS-PAGE gels and blotted onto a polyvinylidene fluoride (PVDF) membrane (Millipore, Bedford, MA). The membranes were incubated with the following specific antibodies: anti-pEGFR (tyr1038), anti-Akt, anti-pAkt (Ser 473), anti-pMET (Y1234/1235), anti-ERK, anti-pERK (T202/Y204), anti-PDGFRβ, anti-ATM, anti-53BP1, anti-pH2AX (S139), anti-pChk2 (T68), and anti-Chk2 (Cell Signaling Technology, Danvers, MA); and anti-EGFR, anti-Met (C-12) (Santa Cruz Biotechnology, Santa Cruz, CA), and anti-GAPDH (Sigma-Aldrich, St. Louis, MO); then, a secondary horseradish peroxidase (HRP)-conjugate antibody was used for immunodetection. The protein bands were visualized using an ECL western blotting kit (Millipore, Bedford, MA) and densitometry analysis with AlphaEaseFC software.

Zhang X., Peng L., Liang Z., Kou Z., Chen Y., Shi G., Li X., Liang Y., Wang F, & Shi Y. (2018). Effects of Aptamer to U87-EGFRvIII Cells on the Proliferation, Radiosensitivity, and Radiotherapy of Glioblastoma Cells. Molecular Therapy. Nucleic Acids, 10, 438-449.

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Immunoprecipitation Protocol for CDC73, ATR, RNAPII

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For immunoprecipitations, cells were lyzed in TX-100 buffer (see under western blotting) containing 100 U/ml Benzonase (Sigma-Aldrich). Lysates were precleared and anti-CDC73 (Bethyl) or anti-pATR T1989 (GeneTex) or anti-RNAPII (F-12, Santa Cruz Biotechnologies) or anti-pCHK2 T68 (used as control antibody, from Cell Signaling) were added. Dynabeads (protein G; Life technologies) were used to isolate antibody-bound complexes.

Landsverk H.B., Sandquist L.E., Sridhara S.C., Rødland G.E., Sabino J.C., de Almeida S.F., Grallert B., Trinkle-Mulcahy L, & Syljuåsen R.G. (2018). Regulation of ATR activity via the RNA polymerase II associated factors CDC73 and PNUTS-PP1. Nucleic Acids Research, 47(4), 1797-1813.

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Immunoblotting for DNA Damage Response

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Antibodies were obtained from the following commercial sources: anti-H2Ax^Ser139 (Millipore; clone JBW301), anti-P53BP1^Ser25 (Novus Biologicals; NB100-1803), anti-53BP1 (ThermoFisher Scientific; PA1-16565), anti-pChk1^S345, anti-pChk2^T68, anti-Chk1, anti-Chk2, anti-ATM (Cell Signaling Technology; 2348S, 2661S, 2G1D5, 1C12 and D2E2 respectively), anti-pATM^S1981 (Rockland, NC9306342), anti-β-actin (Life Technologies, clone AC-15) and anti-vinculin (Abcam, clone ab18058).

Sears C.R., Cooney S.A., Chin-Sinex H., Mendonca M.S, & Turchi J.J. (2016). DNA damage response (DDR) pathway engagement in cisplatin radiosensitization of non-small cell lung cancer. DNA repair, 40, 35-46.

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Chk1/Chk2 Signaling Pathway Analysis

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Cells were collected using a scraper, washed with PBS and resuspended in lysis buffer (25 mM Tris-HCl pH 7.5, 150 mM NaCl, 1% NP-40, 1 mM EDTA, 1 mM EGTA) containing protease and phosphatase inhibitor cocktail (Roche Molecular Biochemicals). The following antibodies with corresponding concentrations were used: anti-PAR (Trevigen, 1:1000), anti-Chk2 (EMD Millipore, 1:1000), anti-pChk2-T68 (Cell Signaling, 1:1000), anti-Chk1 (Santa Cruz, 1:750), anti-pChk1-S345 (Cell Signaling, 1:1000), anti-pKAP1-S824 (Bethyl Laboratories, 1:2000), anti-Cas9 (Cell Signaling Technology, 1:1000), anti-Clathrin Heavy Chain (BD Transduction Laboratories, 1:10 000) and anti-Tubulin (Sigma, 1:10 000).

Vriend L.E., Prakash R., Chen C.C., Vanoli F., Cavallo F., Zhang Y., Jasin M, & Krawczyk P.M. (2016). Distinct genetic control of homologous recombination repair of Cas9-induced double-strand breaks, nicks and paired nicks. Nucleic Acids Research, 44(11), 5204-5217.

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