53BP1 [p Ser25] Antibody

Novus Biologicals | Catalog # NB100-1803

Novus Biologicals
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Key Product Details

Validated by

Independent Antibodies, Biological Validation

Species Reactivity

Validated:

Human, Mouse

Cited:

Human, Mouse

Applications

Validated:

Immunohistochemistry, Immunohistochemistry-Paraffin, Western Blot, Flow Cytometry, Immunocytochemistry/ Immunofluorescence, Immunoprecipitation

Cited:

Western Blot, Immunocytochemistry/ Immunofluorescence

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG
View all 53BP1 Primary Antibodies »

Product Specifications

Immunogen

53BP1 [p Ser25] Antibody was made to a phosphorylated synthetic peptide, which represented a portion of human p53 Binding Protein 1 (GeneID 7158) around serine 25 according to the numbering given in entry NP_005648.1.

Modification

p Ser25

Marker

DNA Double Strand Break Marker

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Description

This antibody can be used as the primary antibody in a PLA assay with the following as complementing antibodies: NB100-97831, NB110-40543, NB100-322, NB200-171, NB100-68265

Scientific Data Images for 53BP1 [p Ser25] Antibody

Western Blot: 53BP1 [p Ser25] Antibody [NB100-1803]

Western Blot: 53BP1 [p Ser25] Antibody [NB100-1803]

Western Blot: 53BP1 [p Ser25] Antibody [NB100-1803] - Samples: Whole cell lysate (30 ug or 15 ug for WB; 1 mg for IP, 20% of IP loaded) from 293T cells that were mock treated or treated with NCS (200 ng/ml; 1h). Affinity purified rabbit anti-Phospho 53BP1 antibody at 0.04 ug/ml (A) and 0.1 ug/ml (B) for WB. 53BP1 was immunoprecipitated using affinity purified rabbit anti-53BP1 Antibody (Catalog #NB100-304). Detection: Chemiluminescence with exposure times of 3 seconds (A and B). Bands appear at ~350 kDa as the observed molecular weight and the theoretical molecular weight is 214 kDa.
Flow Cytometry: 53BP1 [p Ser25] Antibody [NB100-1803]

Flow Cytometry: 53BP1 [p Ser25] Antibody [NB100-1803]

Flow Cytometry: 53BP1 [p Ser25] Antibody [NB100-1803] - Flow cytometric analysis of phospho 53BP1 (S25). Jurkat cells were treated with neocarzinostatin (NCS) for 1.5 hrs. 1 Million cells were fixed, permeabilized, and stained with 1.5 ug/ml antiphospho 53BP1 (S25) (Catalog #NB100-1803) in a 150 ul reaction. Isotype control (black), untreated (red), NCS treated (blue).
Immunohistochemistry-Paraffin: 53BP1 [p Ser25] Antibody [NB100-1803]

Immunohistochemistry-Paraffin: 53BP1 [p Ser25] Antibody [NB100-1803]

Immunohistochemistry-Paraffin: 53BP1 [p Ser25] Antibody [NB100-1803] - FFPE sections of human stomach carcinoma (left) and mouse teratoma (right). Antibody: Affinity purified rabbit anti- Phospho 53BP1 (S25) used at a dilution of 1:1,000 (1ug/ml). Detection: DAB.

Applications for 53BP1 [p Ser25] Antibody

Application
Recommended Usage

Flow Cytometry

25 ug per 1 million cells in a 150 mcl reaction

Immunocytochemistry/ Immunofluorescence

1:500 - 1:2500

Immunohistochemistry

1:1000 - 1:5000

Immunohistochemistry-Paraffin

1:1000 - 1:5000

Immunoprecipitation

1-4 ug/mg of lysate

Western Blot

1:2000-1:10000
Application Notes
Epitope retrieval with citrate buffer pH 6.0 is recommended for FFPE tissue sections. ICC/IF reactivity reported in (PMID: 27713178).

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Flow Cytometry

Formulation, Preparation, and Storage

Purification

Immunogen affinity purified

Formulation

TBS and 0.1% BSA

Preservative

0.09% Sodium Azide

Concentration

0.2 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C. Do not freeze.

Background: 53BP1

Tumor protein p53 binding protein 1 (P53-binding protein 1 or 53BP1) plays a critical role in tumor suppression and is a putative substrate of ATM kinase with a theoretical molecular weight of 214 kDa. Upon DNA damage, it is phosphorylated and relocalizes to the presumptive sites of damage, specifically, double-strand breaks. 53BP1 plays a key role in response to DNA damage, acts as a signaling checkpoint during mitosis, and enhances TP53-mediated transcriptional activation. Originally identified as p53's transcriptional enhancing partner, 53BP1 is known as a key substrate for ataxia telangiectasia mutated (ATM) signaling, whose function to generate gamma H2AX may be partially compensated by the activity of DNA-dependent kinase (DNA-PK). 53BP1 relocalizes to discrete foci overlapping with gamma phosphorylated histone H2AX; demarcating DNA double strand breaks (DSBs) sites following exposure to radiation (1). 53BP1 functions downstream of gamma H2AX-dependent proteins that collectively establish ionizing radiation induced foci at DSBs. 53BP1 is downstream of Mre11/Rad50/NBS1 (MRN complex), MDC1, RNF8, RNF168 and HERC2 which recruit 53BP1 to the DSB site, suggesting a role in DNA repair through genomic stability maintenance (2).

References

1.Henry, E., Souissi-Sahraoui, I., Deynoux, M., Lefevre, A., Barroca, V., Campalans, A.,... Arcangeli, M. L. (2019). Human hematopoietic stem/progenitor cells display ROS-dependent long-term hematopoietic defects after exposure to low dose of ionizing radiations. Haematologica. doi:10.3324/haematol.2019.226936

2.Janoshazi, A. K., Horton, J. K., Zhao, M. L., Prasad, R., Scappini, E. L., Tucker, C. J., & Wilson, S. H. (2020). Shining light on the response to repair intermediates in DNA of living cells. DNA Repair (Amst), 85, 102749. doi:10.1016/j.dnarep.2019.102749

Long Name

p53 Binding Protein 1

Alternate Names

p202, TP53BP1

Entrez Gene IDs

7158 (Human)

Gene Symbol

TP53BP1

UniProt

Q12888

Additional 53BP1 Products

Product Documents for 53BP1 [p Ser25] Antibody

Certificate of Analysis

To download a Certificate of Analysis, please enter a lot or batch number in the search box below.

Download SDS

Product Specific Notices for 53BP1 [p Ser25] Antibody

This antibody can be used as the primary antibody in a PLA assay with the following as complementing antibodies: NB100-97831, NB110-40543, NB100-322, NB200-171, NB100-68265

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.

Related Research Areas

p53 Pathway

Citations for 53BP1 [p Ser25] Antibody

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Protocols

View specific protocols for 53BP1 [p Ser25] Antibody (NB100-1803):

Flow Cytometry Protocol for 53BP1 Antibody (NB100-1803):
Flow Cytometry Protocol for Indirect Intracellular Staining of Cultured Cells Grown in Suspension:
Reagents Required:

16% parafolmaldehyde (PFA) (Electron Microsopy Sciences RT 15710)
90% Methanol (ice cold)
PBS (Phosphate Buffered Saline pH 7.2)
11.6 g sodium chloride
100 ml 1 M phosphate buffer
0.26 Ml 4 M sodium hydroxide
DI H2O to 1 liter
ICSB (IntraCellular Staining Buffer)
495 ml PBS
5 ml FBS (feteal bovine serum)
0.36 ml 9% soldiuim azide
12 x 75 FACS tubes (BD Falcon #352054)


Cell Fixation and Permeabilization:


1. For cells grown in suspension, add fresh 16% PFA to achieve a final concentration of 1.54% directly to the cells in media (1-2 x 106 cell/ml).
2. Fix cells at room temperature (RT) for 10 minutes.
3. Incubate and fix on ice for an additional 30 minutes.
4. Spin fixed cells at 250 x g for 5 minutes at 20C.
5. Pour off media + PFA into dedicated PFA waste containier.
6. Permeabilize cells by resuspending in ice cold 90% methanol to a final cell concentration of 1 X 106 cells/ml.
7. Cells may be stored at -70 to -20C for up to a month.


Intracellular Staining:


1. Aliquot 1 ml (1 x 106 cells) of cells in methanol for each tube/sample.
2. Spin at 250 x g for 5 minutes at 20C, break set to slow (perform all subsequent spins at these conditions)
3. Pour off methanol.
4. Resuspend cells in 1 ml ICSB for wash.
5. Pour off wash and blot tube on paper towels.
6. Resuspend cells in 50 mcl of primary antibody at desired dilution.
7. Incubate at RT for 1 hour.
8. Add 1 ml of ICSB.
9. Spin.
10. Pour off and blot tube.
11. Resuspend in 100 mcl of conjugated secondary antibody at desired dilution.
12. Incubate 30 minutes at RT in dark.
13. Add 1 ml ICSB for 2nd wash.
14. Spin, pour off, and blot tube.
15. Resuspend in 100 mcl ICSB.
16. Analyze on a flow cytometer.


Notes:

This is a recommended protocol, and additional optimization may be required for any individual experiment.

Find general support by application which include: protocols, troubleshooting, illustrated assays, videos and webinars.

FAQs for 53BP1 [p Ser25] Antibody

Showing  1 - 3 of 3 FAQs Showing All

  • Q: Hello I want to ask you for an advice in case of antibodies. In our research we focus on DNA repair foci and use your Novus antibodies for phosphorylated histone H2AX (monoclonal mouse) and protein 53BP1(polyclonal rabbit). Now we want to use antobody for phosphorylated 53BP1 protein to increase specifity and I want to ask you which from your NOVUS antibodies will be the best for us.

    A:

    We currently stock just one antibody to phosphorylated 53BP1, which you can see at the following link: NB100-1803. This is guaranteed for detection of 53BP1 (pSer25) in human and mouse samples by WB, FLOW, ICC/IF,IHC-P, IP and PLA. As this antibody is a rabbit polyclonal, you would be unable to stain for both 53BP1 and its phosphorylated isoform in the same sample, unless you used an antibody from a different host for detection of total 53BP1 or used directly conjugated primaries. Our mouse monoclonal to 53BP1, with catalogue number NBP2-25028, is validated for detection of the human protein by WB, FLOW and IHC. Unfortunately we do not currently stock any other antibodies to phosphorylated 53BP1.

  • Q: I would like to know if this ab would also work for phosphorylated 53BP1? Or if you have another ab that would detect it?

    A: NB100-904 is a cocktail of NB100-304 and NB100-305. These two antibodies recognize between residues 350 and 400 and residues 1925 and the C-terminus (residue 1972), respectively. There are multiple phosphorylations between residues 350 and 400 and as you can see from our datasheet, NB100-304 recognizes 53BP1. We do sell one phosphospecific antibody to 53BP1, catalog number NB100-1803.

  • Q: We're looking for anti53BP1 antibody covalently labeled with a fluorofor preferabley Cy3. Do you make it?

    A:

    We sell 53BP1 antibodies available conjugated to DyLight 488, 550 (very similar in spectrum to Cy3) and 650. You may also purchase the unlabeled antibodies and conjugate them to the fluorophores of your choice. The catalog numbers that you may be interested in are: NB100-904R, NB100-304R, NB100-305R.

  • Q: Hello I want to ask you for an advice in case of antibodies. In our research we focus on DNA repair foci and use your Novus antibodies for phosphorylated histone H2AX (monoclonal mouse) and protein 53BP1(polyclonal rabbit). Now we want to use antobody for phosphorylated 53BP1 protein to increase specifity and I want to ask you which from your NOVUS antibodies will be the best for us.

    A:

    We currently stock just one antibody to phosphorylated 53BP1, which you can see at the following link: NB100-1803. This is guaranteed for detection of 53BP1 (pSer25) in human and mouse samples by WB, FLOW, ICC/IF,IHC-P, IP and PLA. As this antibody is a rabbit polyclonal, you would be unable to stain for both 53BP1 and its phosphorylated isoform in the same sample, unless you used an antibody from a different host for detection of total 53BP1 or used directly conjugated primaries. Our mouse monoclonal to 53BP1, with catalogue number NBP2-25028, is validated for detection of the human protein by WB, FLOW and IHC. Unfortunately we do not currently stock any other antibodies to phosphorylated 53BP1.

  • Q: I would like to know if this ab would also work for phosphorylated 53BP1? Or if you have another ab that would detect it?

    A: NB100-904 is a cocktail of NB100-304 and NB100-305. These two antibodies recognize between residues 350 and 400 and residues 1925 and the C-terminus (residue 1972), respectively. There are multiple phosphorylations between residues 350 and 400 and as you can see from our datasheet, NB100-304 recognizes 53BP1. We do sell one phosphospecific antibody to 53BP1, catalog number NB100-1803.

  • Q: We're looking for anti53BP1 antibody covalently labeled with a fluorofor preferabley Cy3. Do you make it?

    A:

    We sell 53BP1 antibodies available conjugated to DyLight 488, 550 (very similar in spectrum to Cy3) and 650. You may also purchase the unlabeled antibodies and conjugate them to the fluorophores of your choice. The catalog numbers that you may be interested in are: NB100-904R, NB100-304R, NB100-305R.

  • Q: Hello I want to ask you for an advice in case of antibodies. In our research we focus on DNA repair foci and use your Novus antibodies for phosphorylated histone H2AX (monoclonal mouse) and protein 53BP1(polyclonal rabbit). Now we want to use antobody for phosphorylated 53BP1 protein to increase specifity and I want to ask you which from your NOVUS antibodies will be the best for us.

    A:

    We currently stock just one antibody to phosphorylated 53BP1, which you can see at the following link: NB100-1803. This is guaranteed for detection of 53BP1 (pSer25) in human and mouse samples by WB, FLOW, ICC/IF,IHC-P, IP and PLA. As this antibody is a rabbit polyclonal, you would be unable to stain for both 53BP1 and its phosphorylated isoform in the same sample, unless you used an antibody from a different host for detection of total 53BP1 or used directly conjugated primaries. Our mouse monoclonal to 53BP1, with catalogue number NBP2-25028, is validated for detection of the human protein by WB, FLOW and IHC. Unfortunately we do not currently stock any other antibodies to phosphorylated 53BP1.

  • Q: I would like to know if this ab would also work for phosphorylated 53BP1? Or if you have another ab that would detect it?

    A: NB100-904 is a cocktail of NB100-304 and NB100-305. These two antibodies recognize between residues 350 and 400 and residues 1925 and the C-terminus (residue 1972), respectively. There are multiple phosphorylations between residues 350 and 400 and as you can see from our datasheet, NB100-304 recognizes 53BP1. We do sell one phosphospecific antibody to 53BP1, catalog number NB100-1803.

  • Q: We're looking for anti53BP1 antibody covalently labeled with a fluorofor preferabley Cy3. Do you make it?

    A:

    We sell 53BP1 antibodies available conjugated to DyLight 488, 550 (very similar in spectrum to Cy3) and 650. You may also purchase the unlabeled antibodies and conjugate them to the fluorophores of your choice. The catalog numbers that you may be interested in are: NB100-904R, NB100-304R, NB100-305R.

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