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Anti nbs1

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Anti-NBS1 is a primary antibody that specifically recognizes the NBS1 (Nibrin) protein. NBS1 is a component of the MRE11-RAD50-NBS1 (MRN) complex, which is involved in DNA double-strand break repair and cell cycle checkpoint activation in response to DNA damage.

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Lab products found in correlation

Anti rad50, by GeneTex (3 mentions) Anti mre11, by GeneTex (3 mentions) Anti phospho atm ser1981, by Abcam (2 mentions) Odyssey system, by LI COR (2 mentions) Anti flag, by Merck Group (2 mentions) Ku55933, by Merck Group (2 mentions) Anti β actin, by Cell Signaling Technology (2 mentions) Anti phospho chk2 thr68, by Cell Signaling Technology (2 mentions) Anti kap1, by Abcam (2 mentions) Anti phospho ser824 kap1, by Fortis Life Sciences (2 mentions) Anti mbp, by Rockland Immunochemicals (2 mentions) Anti chk2, by GeneTex (2 mentions) Anti phospho ser15 p53, by Abcam (2 mentions) Protease and phosphatase inhibitors, by GoldBio (1 mentions) Anti vinculin, by Merck Group (1 mentions) Anti β actin, by Novus Biologicals (1 mentions) Anti mre11, by Cell Signaling Technology (1 mentions) Anti fancd2, by Novus Biologicals (1 mentions) Anti gapdh, by Novus Biologicals (1 mentions) Anti lamin b1, by Thermo Fisher Scientific (1 mentions)

5 protocols using anti nbs1

1

ATM/DNA-PK Regulation of DDR Signaling

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HCT116 Flp-In T-REx cells were treated with 10 μg/mL doxycycline (Dox) for 16 hr to induce the expression of homeodomain proteins. KU-55933 (EMD Millipore) and NU-7441 (R&D Pharmaceuticals; 3712) were used to treat cells for 1 hr for ATM or DNA-PK inhibition, respectively. DNA-damaging agents and hydrogen peroxide treatments were used to treat cells as indicated in figure legends. After treatment, cells were scraped into PBS and centrifuged at 800 × g for 2 min. Cells were lysed with cell lysis buffer, and lysates were clarified at 10,000 × g for 10 min. The protein concentrations of the lysates were quantified using Bradford assay, and protein levels were normalized for all samples. Primary antibodies used were anti-ATM (Santa Cruz), anti-phospho-Ser1981-ATM (Abcam), anti-Kap1 (Abcam; ab22553), anti-phospho-Ser824-Kap1 (Bethyl Laboratories; A300–767A), anti-Nbs1 (Genetex; GTX70224), anti-Rad50 (Genetex; GTX70228), anti-Mre11 (Genetex; GTX70212), anti-MBP (Rockland; 200–401–385), anti-Flag (Sigma; A1205), anti-β-actin (Cell Signaling; 4970), anti-Chk2 (Genetex; GTX70295), anti-phospho-Thr68-Chk2 (Cell Signaling; 2661S), anti-phospho-Ser15-p53 (Abcam; ab1431), and anti-phospho-Ser15-p53 (mouse) (Assay Biotech; A7180). Alexa Fluor 68-anti-rabbit and IRDye 800 anti-mouse were the secondary antibodies used. Membranes were scanned and quantified using the Odyssey system (Li-Cor).
Johnson T.E., Lee J.H., Myler L.R., Zhou Y., Mosley T.J., Yang S.H., Uprety N., Kim J, & Paull T.T. (2018). Homeodomain Proteins Directly Regulate ATM Kinase Activity. Cell reports, 24(6), 1471-1483.
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2

Western Blot Analysis of DNA Repair Proteins

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Cells were collected by trypsinization and lysed in RIPA lysis buffer (150 mM NaCl, 1.0% IGEPAL CA-630, 0.5% sodium deoxycholate, 0.1% SDS, 50 mM Tris base, pH 8.0). Phosphatase and protease inhibitors (Gold Biotechnology) were added freshly to the lysis buffer. Following gel electrophoresis and transfer of cell extracts onto nitrocellulose, membranes were incubated for 1 hr or overnight in blocking buffer (5% milk in TBS + 0.1% tween). Membranes were subsequently incubated with primary antibodies diluted in antibody buffer (3% BSA in TBS + 0.1% tween) for 2 hrs at room temperature or overnight at 4°C. Detection was achieved using appropriate horseradish peroxidase (HRP)-conjugated secondary antibodies. Anti-ZRANB3 (1:5,000, Bethyl Laboratories), anti-SMARCAL1 (1:1,000, Santa Cruz Biotechnology), anti-BRCA1 (1:500, Bethyl Laboratories), anti-BRCA2 (1:1,000, Bethyl Laboratories), anti-vinculin (1:100,000, Sigma-Aldrich), anti-β-actin (1:100,000, Novus Biologicals), anti-GAPDH (1:5,000, Novus Biologicals), anti-HLTF (1:2,000, Abcam), anti-LAMIN B1 (Thermo Fisher Scientific), anti-FANCD2 (1:1,000, Novus Biologicals), anti-PCNA (1:100,000, Abcam), anti-NBS1 (GeneTex), anti-MRE11 (Cell Signaling Technology), anti-RPA2 (Bethyl Laboratories) antibodies were used in western blot experiments.
Taglialatela A., Alvarez S., Leuzzi G., Sannino V., Ranjha L., Huang J.W., Madubata C., Anand R., Levy B., Rabadan R., Cejka P., Costanzo V, & Ciccia A. (2017). Restoration of replication fork stability in BRCA1- and BRCA2-deficient cells by inactivation of SNF2-family fork remodelers. Molecular cell, 68(2), 414-430.e8.
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3

ATM/DNA-PK Regulation of DDR Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols
HCT116 Flp-In T-REx cells were treated with 10 μg/mL doxycycline (Dox) for 16 hr to induce the expression of homeodomain proteins. KU-55933 (EMD Millipore) and NU-7441 (R&D Pharmaceuticals; 3712) were used to treat cells for 1 hr for ATM or DNA-PK inhibition, respectively. DNA-damaging agents and hydrogen peroxide treatments were used to treat cells as indicated in figure legends. After treatment, cells were scraped into PBS and centrifuged at 800 × g for 2 min. Cells were lysed with cell lysis buffer, and lysates were clarified at 10,000 × g for 10 min. The protein concentrations of the lysates were quantified using Bradford assay, and protein levels were normalized for all samples. Primary antibodies used were anti-ATM (Santa Cruz), anti-phospho-Ser1981-ATM (Abcam), anti-Kap1 (Abcam; ab22553), anti-phospho-Ser824-Kap1 (Bethyl Laboratories; A300–767A), anti-Nbs1 (Genetex; GTX70224), anti-Rad50 (Genetex; GTX70228), anti-Mre11 (Genetex; GTX70212), anti-MBP (Rockland; 200–401–385), anti-Flag (Sigma; A1205), anti-β-actin (Cell Signaling; 4970), anti-Chk2 (Genetex; GTX70295), anti-phospho-Thr68-Chk2 (Cell Signaling; 2661S), anti-phospho-Ser15-p53 (Abcam; ab1431), and anti-phospho-Ser15-p53 (mouse) (Assay Biotech; A7180). Alexa Fluor 68-anti-rabbit and IRDye 800 anti-mouse were the secondary antibodies used. Membranes were scanned and quantified using the Odyssey system (Li-Cor).
Johnson T.E., Lee J.H., Myler L.R., Zhou Y., Mosley T.J., Yang S.H., Uprety N., Kim J, & Paull T.T. (2018). Homeodomain Proteins Directly Regulate ATM Kinase Activity. Cell reports, 24(6), 1471-1483.
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4

Western Blot Analysis of DNA Repair Proteins

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Western blots were performed with primary antibodies anti-RAD50 (GTX70228, Genetex, Irvine, CA, USA), anti-MRE11 (GTX70212, Genetex, Irvine, CA, USA), anti-NBS1 (GTX70224, Genetex, Irvine, CA, USA), anti-6xHIS (#12698, Cell Signaling Technology, Danvers, MA, USA), anti-α-tubulin (GTX112141, Genetex, Irvine, CA, USA), and anti-P84 (GTX70220, Genetex, Irvine, CA, USA), as described previously [36 (link),37 (link)].
Ko J.M., Lam S.Y., Ning L., Chai A.W., Lei L.C., Choi S.S., Wong C.W, & Lung M.L. (2021). RAD50 Loss of Function Variants in the Zinc Hook Domain Associated with Higher Risk of Familial Esophageal Squamous Cell Carcinoma. Cancers, 13(18), 4715.
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5

Protein Expression Analysis Protocol

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Exponentiall growing cells were collected by trypsinization, washed with PBS, and lysed in RIPA buffer (50 mM Tris–HCl, pH 7.2, 150 mM NaCl, 1% Nonidet P-40, 1% Sodium deoxycholate and 0.1% Sodium dodecylsulfate) containing protease inhibitor cocktail (Roche Japan, Tokyo, Japan). Protein concentrations were determined by BCA protein assay (Thermo Fisher Scientific), and 8 µg of proteins were electrophoresed on 5–10% SDS–polyacrylamide gels, and then electrophoretically transferred to a polyvinyl difluoride membrane. Primary antibodies used in this study are as follows: anti-RAD51 (GTX70230, GeneTex), anti-MRE11 (clone 12D7, GeneTex), anti-ATM (clone 1A1, GeneTex), anti-Rad50 (clone 13B3, GeneTex), anti-NBS1 (clone 1D7, GeneTex), anti-BRCA1 (clone 6B4, GeneTex), anti-BRCA2 (clone 1B8, GeneTex), anti-Ku86 (clone 111, Kamiya Biochemical co., Seattle, WA, USA), anti-Ku70 (clone N3H10, Kamiya Biochemical co.), anti-DNA-PKcs (MC-365, Kamiya Biochemical co.), anti-XRCC4 (GTX70293, GeneTex), anti-Lig IV (GTX74360, GeneTex), anti-XLF (A300-730, BETHYL Laboratories, Inc.), and anti-γ-tubulin (clone DTU-88, Sigma-Aldrich).
Tanaka K., Suzuki K., Miyashita K., Wakasa K., Kawano M., Nakatsu Y., Tsumura H., Yoshida M.A, & Oda S. (2022). Activation of recombinational repair in Ewing sarcoma cells carrying EWS-FLI1 fusion gene by chromosome translocation. Scientific Reports, 12, 14764.
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