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Anti 53bp1

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Anti-53BP1 is a primary antibody that recognizes the 53BP1 protein. 53BP1 is a DNA damage response protein involved in the repair of double-strand breaks in DNA.

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Lab products found in correlation

Alexa fluor 488, by Thermo Fisher Scientific (3 mentions) Anti γh2ax, by Abcam (2 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions) Anti γ h2ax mouse monoclonal antibody, by Merck Group (2 mentions) Alexa fluor 555, by Thermo Fisher Scientific (2 mentions) Fluoroshield with dapi, by Merck Group (2 mentions) Alexa 488 555 conjugated secondary antibody, by Thermo Fisher Scientific (2 mentions) Anti pcna rabbit polyclonal antibody, by Abcam (2 mentions) Anti γh2ax, by Merck Group (2 mentions) Anti γh2ax, by Cell Signaling Technology (2 mentions) Anti phospho histone h2a x ser139, by Merck Group (1 mentions) Anti rad51, by Abcam (1 mentions) Anti rnf168, by Abcam (1 mentions) Anti cxcl5, by Abcam (1 mentions) Anti g0s2, by Proteintech (1 mentions) Mir nc, by GenePharma (1 mentions) Inhibitor nc, by GenePharma (1 mentions) Anti lig4 12695 1 ap, by Proteintech (1 mentions) Mir 1246 inhibitor, by GenePharma (1 mentions) Anti tsg101, by Abcam (1 mentions)

Spelling variants of this product

Rabbit polyclonal anti-mouse 53BP1 antibody (2 citations) Rabbit anti-53BP1 antibody (2 citations) Anti-53BP1 (ab175933, (2 citations) Rabbit anti-53BP1 polyclonal antibody (2 citations) Rabbit anti-53BP1 (2 citations) Anti-53BP1 antibody (2 citations)

19 protocols using anti 53bp1

1

Antibody Immunostaining Protocol

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The following antibodies were used in this study: an anti-G0S2 (dilution 1:200; Proteintech, IL, USA); an anti-phospho-Histone H2A.X (Ser139) (dilution1:1000; EMD Millipore, Billerica, MA, USA); anti-53BP1 (dilution1:500), anti-RNF168 (dilution1:500), anti-CXCL5 (dilution 1:1000) and anti-Rad51 (dilution 1:1000)(Abcam, Cambridge, MA, USA). The secondary antibodies were from Jackson ImmunoResearch Laboratories (West Grove, PA, USA). Cell culture media and other reagents were from Invitrogen (Carlsbad, CA, USA), Sigma-Aldrich (St. Louis, MO, USA) or Peprotech (Rocky Hill, NJ, USA).
Wang Y., Hou Y., Zhang W., Alvarez A.A., Bai Y., Hu B., Cheng S.Y., Yang K., Li Y, & Feng H. (2019). Lipolytic inhibitor G0S2 modulates glioma stem-like cell radiation response. Journal of Experimental & Clinical Cancer Research : CR, 38, 147.
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2

miR-1246 Modulates NHEJ Repair Efficiency

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EJ5-GFP plasmids were linearised by HindIII enzyme digestion and pCherry vectors were used to detect the
double-strand break (DSB) repair efficiency of nonhomologous end joining
(NHEJ).33 –35 (link) miR-1246 mimic (sense:
5′-AAUGGAUUUUUGGAGCAGG-3′, antisense: 5′-UGCUCCAAAAAUCCAUUUU-3′); miR-NC (sense:
5′-UUCUCCGAACGUGUCACGUTT-3′, antisense: 5′-ACGUGACACGUUCGGAGAATT-3′); miR-1246
inhibitor (5′-CCUGCUCCAAAAAUCCAUU-3′); and inhibitor-NC
(5′-CAGUACUUUUGUGUAGUAGUACAA-3′) were purchased from GenePharma (Shanghai,
China).
Antibodies used in this study were as follows: anti-53BP1 (Abcam,
Cambridge, UK), anti-LIG4 (12695-1-AP; Proteintech Group Inc., Rosemont, IL, USA),
anti-CD63 (H-193, sc-15363; Santa Cruz Biotechnology, Santa Cruz, CA, USA),
anti-TSG101 (ab133586; Abcam, Cambridge, MA, USA), anti-GAPDH (TA309157; Beijing
Zhongshan Jinqiao Biotechnology Co., Ltd., Beijing, China), and anti-β-actin
(TA-09; Beijing Zhongshan Jinqiao Biotechnology Co., Ltd.).
Mo L.J., Song M., Huang Q.H., Guan H., Liu X.D., Xie D.F., Huang B., Huang R.X, & Zhou P.K. (2018). Exosome-packaged miR-1246 contributes to bystander DNA damage by targeting LIG4. British Journal of Cancer, 119(4), 492-502.
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3

Antibody Protocols for Centrosomal Proteins

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Antibodies against Cep192 (1–211 aa; used at 0.5 µg/ml), SAS-6 (501–657 aa; used at 0.5 µg/ml), and Plk4 (501–657 aa; used at 1 µg/ml) were previously described (Wong et al., 2015 (link)). The following antibodies were purchased from commercial sources, with their working concentrations indicated in parentheses: anti-Trim37 (1:2,000 for Western blotting; A301-174A; Bethyl Laboratories, Inc.), anti-Usp28 (1:1,000 for Western blot; ab126604; Abcam), anti-Usp28 (1:100 for immunofluorescence; HPA006778; Sigma-Aldrich), anti-53BP1 (1:5,000), anti-Cep152 (1:2,000; Abcam), anti-Cdk5rap2 (1:4,000; Abcam), GTU-88 (anti–γ-tubulin; 1:1,000; Sigma-Aldrich), anti-pericentrin (1 µg/ml; Abcam), anti-CPAP (1:400; Proteintech), anti-p53 (1:100 for Western blot; OP43; EMD Millipore), anti-p53 (1:500 for immunofluorescence; OP140; EMD Millipore), anti-p21 (1:1,000; #2947; Cell Signaling Technology), DM1A (anti–α-tubulin; 1:5,000; Sigma-Aldrich), anti-GAPDH (1:1,000; 14C10; Cell Signaling Technology), PCM-1 (1:400; #5259; Cell Signaling Technology), and Fab fragment (goat anti–rabbit IgG; 30 µg/ml; Jackson ImmunoResearch Laboratories, Inc.). Secondary antibodies were purchased from Jackson ImmunoResearch Laboratories, Inc.
Meitinger F., Anzola J.V., Kaulich M., Richardson A., Stender J.D., Benner C., Glass C.K., Dowdy S.F., Desai A., Shiau A.K, & Oegema K. (2016). 53BP1 and USP28 mediate p53 activation and G1 arrest after centrosome loss or extended mitotic duration. The Journal of Cell Biology, 214(2), 155-166.
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4

Comprehensive Antibody Panel for Immune Signaling

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The following antibodies were used: anti–PD-L1 (Abcam, no. ab213480), anti–Caspase-3 (CST, no. 9662), anti-GSDME (Abcam, no. ab215191), anti-Granzyme B (Abcam, no. ab255598), anti-STING (CST, no. 50494), anti–phospho-STING (Ser365) (CST, no. 72971), anti–TBK-1 (CST, no. 3504), anti–phospho-TBK-1 (Ser172) (CST, no. 5483), anti–IRF-3 (CST, no. 4302), anti–phospho-IRF3 (Ser396) (CST, no. 4947S), anti–γ-H2AX (phospho Ser139) (Abcam, no. ab81299), anti-53BP1 (Abcam, no. ab175933), anti–phospho-53BP1 (Ser25) (Abcam, no. ab70323), anti–TREX-1 (Santa Cruz, no. sc-133112), anti-GFP (Beyotime Inc., no. AG279), and anti–β-actin (Abcam, no. ab8226). For flow cytometry analysis, anti–CD3-APC (Biolegend, no. 100236) and anti–CD8-PE (Biolegend, no. 100707) were used.
Shi X., Yang Y., Zhang W., Wang J., Xiao D., Ren H., Wang T., Gao F., Liu Z., Zhou K., Li P., Zhou Z., Zhang P., Shen X., Liu Y., Zhao J., Wang Z., Liu F., Shao C., Wu D, & Zhang H. (2022). FLASH X-ray spares intestinal crypts from pyroptosis initiated by cGAS-STING activation upon radioimmunotherapy. Proceedings of the National Academy of Sciences of the United States of America, 119(43), e2208506119.
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5

Comprehensive Antibody Catalog for Research

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In this work, the following antibodies were used: primary antibodies: anti-hnRNP UL1 (Abcam #ab68480 or Santa Cruz Biotechnology #sc-393434), anti-FLAG (Sigma #A8592), anti-Actin (MP #691001), anti-Fibrillarin (Santa Cruz Biotechnology #sc-25397), anti-Nucleolin (Abcam #ab22758), anti-FUS (Santa Cruz Biotechnology #sc-47711), anti-γH2A.X (Santa Cruz Biotechnology #sc-517348), anti-RPS6 (Abcam #ab70227), anti-RPS15 (Antikoerper #ABIN2786563), anti-RPA32 (Bethyl Laboratories #A300-245A), anti-pChk1 (Cell Signaling Technology #2341), anti-XRCC1 (Invitrogen #MA5-13412), anti-53BP1 (Abcam #ab175933), anti-Rad50 (Abcam #ab124682), anti-RPA194 (Santa Cruz Biotechnology #sc-46699), normal mouse IgG (Santa Cruz Biotechnology #sc-2025), and anti-digoxygenin-AP Fab fragments (Roche #11093274910); secondary antibodies: goat anti-mouse IgG-horseradish peroxidase (HRP) (Santa Cruz Biotechnology #sc-516102), goat anti-rabbit IgG-HRP (Santa Cruz Biotechnology #sc-2004), anti-mouse Alexa Fluor 555 (Thermo Fisher Scientific #A21422), anti-mouse Alexa Fluor 488 (Thermo Fisher Scientific #A32723), anti-rabbit Alexa Fluor 555 (Thermo Fisher Scientific #A32732) or anti-rabbit Alexa Fluor 488 (Thermo Fisher Scientific #A32731).
Cichocka M., Karlik A., Plewka P., Gawade K., Stępień A., Świergiel P., Gadgil A, & Raczyńska K.D. (2022). The Novel Role of hnRNP UL1 in Human Cell Nucleoli. International Journal of Biological Sciences, 18(13), 4809-4823.
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6

Immunofluorescence Analysis of DNA Damage Response

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48 hours after siRNA transfection, cells were harvested, reseeded in chamber slides and irradiated 4 hours later. At indicated times after IR, cells were washed, fixed, permeabilized and incubated with blocking buffer. Primary antibodies anti-RAD51 (Calbiochem, PC130-100UL), anti-cyclin B1 (EMD Millipore, 05-373) and anti-53BP1 (Abcam, ab21083) were added to the cells and incubated overnight. Secondary antibodies anti-mouse Alexa Fluor 488 (Invitrogen, A11017) and anti-rabbit Cyanine Cy3 (Jackson, 111-165-008) were applied together with 1 µg/ml DAPI (Invitrogen) nuclear counterstain for 1,5 hours at room temperature. Slides were examined on the fluorescent microscope imager Z1 (Zeiss).
Niedermaier B., Sak A., Zernickel E., Xu S., Groneberg M, & Stuschke M. (2019). Targeting ARID1A-mutant colorectal cancer: depletion of ARID1B increases radiosensitivity and modulates DNA damage response. Scientific Reports, 9, 18207.
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7

Immunofluorescence Quantification of DNA Damage

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After irradiation, the cells on the TW insert membrane (#3460, Corning) were fixed with 4% paraformaldehyde for 15 min, washed with PBS, permeabilized (100 mM TrisCl pH 7.4, 50 mM EDTA, 0.5% Triton100 in H2O) and blocked (3% BSA, 0.1% Tween20, 4xSSC, 7.7 mM NaN3 in H2O). The cells on the membranes were incubated overnight at 4 °C with the primary antibodies anti-phospho histone γH2AX (Millipore, 05-636) and anti-53BP1 (Abcam, ab21083) in PBS. The cells were washed with PBS and incubated with a secondary antibody labeled with Alexa488 (A 2102, Invitrogen) or Cy3 (BA 1034-05, Boster) and DAPI 1 mg/ml (Thermo Scientific) for 90 min at room temperature. The membranes were cut manually and transferred onto slides, mounted with immu-mount and cover-slipped. The γH2AX and 53PB1 foci per nucleus were identified by eye and calculated manually using a Leica DMi8 fluorescent microscope (USA).
Bannik K., Madas B., Jarzombek M., Sutter A., Siemeister G., Mumberg D, & Zitzmann-Kolbe S. (2019). Radiobiological effects of the alpha emitter Ra-223 on tumor cells. Scientific Reports, 9, 18489.
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8

Immunofluorescent Analysis of Muscle Cells

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Cells were first washed with Phosphate Buffer Saline (Thermo Fisher) and fixed with 4% Paraformaldehyde (MS). Cells were stained with the following primary antibodies and concentrations, MYHC‐IIb eFluor 660 (50‐6503‐32; Thermo Fisher; 1:100), α‐actinin (sc‐7453; Santa Cruz; 1:500), and myogenin (sc‐576; Santa Cruz; 1:200). Anti‐PARP‐1 (Santa, SC‐8007), anti‐XRCC1 (Cell Signalling, 2735S), anti‐gamma H2AX (Abcam, 2893), anti‐53BP1 (Abcam, ab175933), and anti‐Osteocalcin (Santa, SC‐365797).The following secondary antibodies were also used together with non‐conjugated primary antibodies, Goat‐anti‐mouse Alexa Fluor 488 (A11001; Thermo Fisher; 1:500), Goat‐anti‐rabbit Alexa Fluor 594 (A11012; Thermo Fisher; 1:500), and Goat‐anti‐mouse Alexa Fluor 647 (A21235; Thermo Fisher; 1:500). 4'6‐Diamidino‐2‐Phenylindole (d9542; Sigma) was used as a nuclear counter stain according to manufacturer's recommendations. Stained cells were imaged with a Zeiss fluorescence microscope.
Wang P., Liu X., Chen Y., Jun‐Hao E.T., Yao Z., Min‐Wen J.C., Yan‐Jiang B.C., Ma S., Ma W., Luo L., Guo L., Song D, & Shyh‐Chang N. (2023). Adult progenitor rejuvenation with embryonic factors. Cell Proliferation, 56(5), e13459.
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9

Immunofluorescence Foci Staining Protocol

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Staining and visualization of DNA damage repair foci was essentially performed as described 52 (link). Cells were plated on chamber slides to achieve subconfluency for fixing. Cells were fixed with 4% PFA, permeabilized with 0.5% TritonX-100, and blocked with 5% goat serum in PBS 0.1% TritonX-100 (all Sigma-Aldrich). Cells were incubated with anti-γ-H2AX mouse monoclonal antibody (Millipore; at 1:500 dilution), anti-53BP1 (Abcam; 1:500), anti-PCNA rabbit polyclonal antibody (Abcam; 1:200), or anti-RAD51 mouse monoclonal antibody followed by incubation with rabbit or mouse Alexa-488/-555 conjugated secondary antibody (Invitrogen) at 1:1,000. All slides were counterstained with DAPI, examined and photographed by fluorescence microscopy (Olympus BX51).
Cryosections from human tissue samples were fixed with 2% PFA for 15 minutes and permeabilized with 0.5% TritonX for 10 minutes afterwards. Following 1 hour blocking at room temperature with 5% goat serum/PBS, slides were incubated overnight with primary antibody in a humid chamber at 4°C. Mouse anti-PCNA was co-incubated with rabbit polyclonal anti-53BP1. Secondary antibody including goat anti-rabbit IgG (Alexa Fluor488, Invitrogen, 1:1000 dilution) and goat anti-mouse IgG (Alexa Fluor555, Invitrogen, 1:1000) was incubated at room temperature for 1 hour. Slides were mounted in Fluoroshield with DAPI (Sigma, F6057).
Liu Q., Gheorghiu L., Drumm M., Clayman R., Eidelman A., Wszolek M.F., Olumi A., Feldman A., Wang M., Marcar L., Citrin D.E., Wu C.L., Benes C.H., Efstathiou J.A, & Willers H. (2018). PARP-1 inhibition with or without ionizing radiation confers reactive oxygen species-mediated cytotoxicity preferentially to cancer cells with mutant TP53. Oncogene, 37(21), 2793-2805.
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10

Quantifying DNA Damage Response Proteins

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Cells were seeded on the glass sheets into 6-well plates. Cells were transfected with siRNAs and then exposed to 4 Gy of irradiation. 30 min later, cells were washed twice in PBS, and then incubated with anti-γH2AX (Millipore, FITC conjugate, Billerica, MA, USA), anti-53BP1 (Abcam) or anti-CHMP4C antibody at a dilution of 1:200 overnight at 4 °C. After washing twice in washing buffer (0.05% Tween-20 in PBS), the cells were incubated with goat anti-rabbit lgG (Abcam, Alexa fluor conjugate) at 37 °C for 1 h. Cells were washed twice with washing buffer and once with PBS. The coverslips were then mounted with mounting medium for fluorescence with DAPI (Vectashield). The image of γH2AX and 53BP1 foci was detected using a Leica DMRXA fluorescent microscope (Leica, Wetzlar, Germany). γH2AX and 53BP1 foci were counted in cells with more than five foci.
Li K., Liu J., Tian M., Gao G., Qi X., Pan Y., Ruan J., Liu C, & Su X. (2015). CHMP4C Disruption Sensitizes the Human Lung Cancer Cells to Irradiation. International Journal of Molecular Sciences, 17(1), 18.
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