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Products:Epigenetics and Nuclear Signaling >> DNA / RNA >> DNA Synthesis >> Topoisomerases
Overview
Product name
Anti-Topoisomerase I antibody - ChIP Grade
See all Topoisomerase I products (7) ...
Description
Rabbit polyclonal to Topoisomerase I - ChIP Grade
Tested applications
ICC/IF, IF, IHC-Fr, WB, IP, ChIPmore details
Cross reactivity
Reacts with
Human
Immunogen
Recombinant fragment, corresponding to amino acids 401 - 600 of Human Topoisomerase I.
Positive control
HeLa cells.
Properties
Form
Liquid
Storage instructions
Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
Purity
Whole antiserum
Primary antibody notes
This product can be used as part of an assay for sumoylation activity. Human Aos 1 + Uba 2 (ab3804), Ubc 9 (ab3803) and Sumo 1 (ab3801) can be used to promote in vitro sumoylation of a sumoylation marker (human Topoisomerase I protein fragment) (ab3828). The reaction products can be detected using our Sumo 1 (ab3819 and ab3824) and Topoisomerase I (ab3825) antibodies. Sumoylation assays are carried out in a final volume of 20µl in reaction conditions (20 mM Hepes pH 7.5, 5mM MgCl2, 2mM ATP). This antibody is equivalent to the antibody used in our Sumoylation Control kits. Sumoylation Protocol: 1. Prepare a suitable purified substrate protein. (For the control, use 2µl Topoisomerase I marker for each reaction.) 2. In each reaction, add 4µl E2 to substrate first, then 2µl Sumo 1, 2µl 10x reaction buffer, 2µl E1. Finally, add H2O to bring up to 20µl. We would recommend adding fresh 2mM ATP to be sure that sufficient energy is supplied. 3. The best reaction concentration of proteins is as following: Aos 1 + Uba 2: 7.5µg/ml. Ubc 9: 50µg/ml. SUMO 1: 50µg/ml. For the control assay we recommend running the assay at 37ºC for 30-60 minutes. 4. Detect the reaction products by Western blot using a suitable antibody. For the control reaction use 1/1000 dilution of the supplied Topoisomerase I antibody. Four sumoylated bands should be seen on the gel for the control reaction. This assay has been shown to work with crude extracts. Be aware that Uba 2 contains his-rich regions which might cross-react with antibodies against the 6x-His epitope tag. During western analysis with anti-6x-His antibodies, Uba 2 at 80 kDa might be shown.
Clonality
Polyclonal
Isotype
IgG
Applications
Show applications key
Our Abpromise guarantee covers the use of ab3825 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ICC/IF: Use at an assay dependent dilution. (PubMed: 20655466)
IF: 1/200 - 1/500.(Frozen sections. Incubate for 60 minutes at 25ºC.)
IHC-Fr: Use at an assay dependent dilution.
WB: 1/1000Detects a band of approximately 92 kDa (predicted molecular weight: 91 kDa).
IP: Use at an assay dependent dilution. (PubMed: 17290216)
ChIP: Use at an assay dependent dilution. (PubMed: 20655466)
Target
Function
The reaction catalyzed by topoisomerases leads to the conversion of one topological isomer of DNA to another.
Involvement in disease
Note=A chromosomal aberration involving TOP1 is found in a form of therapy-related myelodysplastic syndrome. Translocation t(11;20)(p15;q11) with NUP98.
Sequence similarities
Belongs to the eukaryotic type I topoisomerase family.
Post-translational
modifications
Sumoylated. Lys-117 is the main site of sumoylation. Sumoylation plays a role in partitioning TOP1 between nucleoli and nucleoplasm. Levels are dramatically increased on camptothecin (CPT) treatment.
Cellular localization
Nucleus > nucleolus. Nucleus > nucleoplasm. Diffuse nuclear localization with some enrichment in nucleoli. On CPT treatment, cleared from nucleoli into nucleoplasm. Sumolyated forms found in both nucleoplasm and nucleoli.
Target information above from: UniProt accessionP11387
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
Alternative names
- DNA topoisomerase 1 antibody
- DNA topoisomerase I antibody
- NUP98 fusion gene antibody
see all
Database links
- Entrez Gene: 7150 Human
- Omim: 126420 Human
- SwissProt: P11387 Human
- Unigene: 472737 Human
- Unigene: 712686 Human
Anti-Topoisomerase I antibody - ChIP Grade images:
Immunocytochemistry/ Immunofluorescence - Topoisomerase I antibody (ab3825)
blue: DAPI; green: Topo I. Hela cell staining.
Western blot - Topoisomerase I antibody (ab3825)
Predicted band size : 91 kDa
Primary antibody incubated at a 1/1000 dilution for 60 minutes at 25ºC.
Western blot - Topoisomerase I antibody (ab3825)
All lanes : Anti-Topoisomerase I antibody - ChIP Grade (ab3825) at 1/5000 dilution
Lane 1 : SW210.5 human SCLC cell xenograft lysate.
Lane 2 : 5M2 human NSCLC cell lysate (bicarb. buffer).
Lane 3 : 5M2 human NSCLC cell lysate (M-Per buffer).
Lane 4 : Normal human lung tissue lysate.
Lane 5 : A431 cell (human epidermoid carcinoma) lysate .
Lane 6 : Rh30 cell (human rhabdomyosarcoma) lysate.
Lysates/proteins at 20 µg per lane.
Secondary
Goat anti rabbit heavy and light (HRP) at 1/25000 dilution
Performed under reducing conditions.
Predicted band size : 91 kDa
This image is courtesy of an Abreview submitted by Mike Campa on 7 September 2005. We do not have any further information relating to this image.
References for Anti-Topoisomerase I antibody - ChIP Grade (ab3825)
This product has been referenced in:
- Biroccio Aet al. DNA damage persistence as determinant of tumor sensitivity to the combination of Topo I inhibitors and telomere-targeting agents. Clin Cancer Res 17:2227-36 (2011). ChIP; Human.Read more (PubMed: 21355072) »
- Kerzendorfer Cet al. Mutations in Cullin 4B result in a human syndrome associated with increased camptothecin-induced topoisomerase I-dependent DNA breaks. Hum Mol Genet 19:1324-34 (2010). WB; Human.Read more (PubMed: 20064923) »
See all 6 publications for this product
Publishing research using ab3825? Please let us know so that we can cite the reference in this datasheet
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"