Overview

  • Product name
    Anti-Thymine Dimer antibody [H3]
  • Description
    Mouse monoclonal [H3] to Thymine Dimer
  • Host species
    Mouse
  • Tested applications
    Suitable for: Southern Blot, ICC, Competitive ELISA, ELISA, ICC/IFmore details
  • Immunogen

    Chemical/ Small Molecule.

  • Positive control
    • ICC/IF: HeLa cells.
  • General notes


    Non-radioactive labeling of DNA is typically based on the enzymatic incorporation of modified nucleotides, carrying a small chemical moiety such as biotin, digoxigenin or fluorescein. These tags are subsequently detected by specific reagents such as streptavidin or a specific antibody coupled to a signal-producing enzyme. Although very efficient and reliable, labeling by in vitro polymerization is time-consuming, expensive, and may require various post-label purification steps to remove an excess of unincorporated precursors. An alternative strategy for DNA labeling, is based on the UV-induced formation of cyclobutane thymine dimers. Several methods have been described for the detection of thymine dimers, which are based on chromato-graphic analysis, and on biochemical analysis with endonucleases specific for UV-irradiated DNA. In addition, methods utilizing antibodies specific for pyrimidine dimers and other UV-induced DNA lesions have evolved, which permit the study of the induction and repair of these lesions without the requirement of in vivo radiolabeling of DNA. Photoimmunodetection, is a rapid, reliable and low-cost supplement to existing methods for nonradioactive DNA labeling. It enables a sensitive and non-radioactive method for labeling, detection, and quantification of high molecular weight (HMW) DNA fragments. The method is based on the introduction of thymine dimers into DNA after separa-tion by pulse field gel electrophoresis (PFGE), followed by detection with thymine dimer specific antibodies. The method does not require any enzymatic or chemical manipulation of the DNA sample. Monoclonal anti-bodies reacting specifically with thymine dimer, facilitate investigations on the apoptotic process and the role of UV-induced pyrimidine dimers in the process of photocarcinogenesis.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
  • Storage buffer
    pH: 7.40
    Preservative: 0.097% Sodium azide
    Constituent: PBS
  • Concentration information loading...
  • Purity
    Protein G purified
  • Primary antibody notes
    Non-radioactive labeling of DNA is typically based on the enzymatic incorporation of modified nucleotides, carrying a small chemical moiety such as biotin, digoxigenin or fluorescein. These tags are subsequently detected by specific reagents such as streptavidin or a specific antibody coupled to a signal-producing enzyme. Although very efficient and reliable, labeling by in vitro polymerization is time-consuming, expensive, and may require various post-label purification steps to remove an excess of unincorporated precursors. An alternative strategy for DNA labeling, is based on the UV-induced formation of cyclobutane thymine dimers. Several methods have been described for the detection of thymine dimers, which are based on chromato-graphic analysis, and on biochemical analysis with endonucleases specific for UV-irradiated DNA. In addition, methods utilizing antibodies specific for pyrimidine dimers and other UV-induced DNA lesions have evolved, which permit the study of the induction and repair of these lesions without the requirement of in vivo radiolabeling of DNA. Photoimmunodetection, is a rapid, reliable and low-cost supplement to existing methods for nonradioactive DNA labeling. It enables a sensitive and non-radioactive method for labeling, detection, and quantification of high molecular weight (HMW) DNA fragments. The method is based on the introduction of thymine dimers into DNA after separa-tion by pulse field gel electrophoresis (PFGE), followed by detection with thymine dimer specific antibodies. The method does not require any enzymatic or chemical manipulation of the DNA sample. Monoclonal anti-bodies reacting specifically with thymine dimer, facilitate investigations on the apoptotic process and the role of UV-induced pyrimidine dimers in the process of photocarcinogenesis.
  • Clonality
    Monoclonal
  • Clone number
    H3
  • Isotype
    IgG1
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab10347 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Southern Blot Use a concentration of 0.5 - 1 µg/ml.
ICC Use at an assay dependent dilution.
Competitive ELISA Use at an assay dependent dilution.
ELISA Use at an assay dependent dilution.
ICC/IF Use at an assay dependent concentration.

Images

  • ab10347 staining Thymine Dimer in HeLa cells by Immunocytochemistry/ Immunofluorescence.
    Cells were fixed in formaldehyde, permabilized using 0.5% Triton X-100, blocked with 5% BSA for 15 minutes at 20°C, then incubated with ab10347 at a 1/250 dilution for 16 hours at 4°C. The secondary used was an Alexa-Fluor 488 conjugated rabbit anti mouse polyclonal, used at a 1/500 dilution.

    See Abreview

References

This product has been referenced in:
  • Torregrosa-Muñumer R  et al. Low doses of ultraviolet radiation and oxidative damage induce dramatic accumulation of mitochondrial DNA replication intermediates, fork regression, and replication initiation shift. Mol Biol Cell 26:4197-208 (2015). Read more (PubMed: 26399294) »
  • Deng Y  et al. A sunblock based on bioadhesive nanoparticles. Nat Mater 14:1278-85 (2015). Read more (PubMed: 26413985) »
See all 7 Publications for this product

Customer reviews and Q&As

1-10 of 19 Abreviews or Q&A

Question
Answer

Apologies, the expiration date should have been 30th September 2012. I have updated our records to reflect the correct date.

Please let me know if there is anything else I can help you with.

Read More
Abcam has not validated the combination of species/application used in this Abreview.
Application
Other
Sample
Human Cell lysate - nuclear (WI38 cell line)

Abcam user community

Verified customer

Submitted Dec 08 2017

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Human skin equivalent model, with and without UVB-)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: pH 8.0 EDTA buffer (90¯C/40min)
Permeabilization
No
Specification
Human skin equivalent model, with and without UVB-
Blocking step
Biocare Medical Background SNIPER as blocking agent for 10 minute(s) · Concentration: 100% · Temperature: 24°C
Fixative
10% NBF

Abcam user community

Verified customer

Submitted Jan 04 2017

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Sample
Human Cell (U2OS)
Specification
U2OS
Permeabilization
Yes - triton-X 100
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Oct 21 2014

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
ELISA
Sample
Human Cell (u2os)
Specification
u2os
Type
Direct
Blocking step
Milk as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 21°C

Dr. Andrew Cobb

Verified customer

Submitted Jul 16 2013

Answer

Thank you for your message.

I can confirm that Thymine Dimer is not a species specific target, and so this antibody should work in pig.

As far as we are aware, ab10347 has never been tested in IHC-P. All tested applications covered by the 6 month guarantee are specified on our individual datasheets, and these are updated as soon as any new information is brought to our attention.

If the customer would like to test ab10347 in IHC-P, please do not hesitate to contact me again prior to the purchase by replying to this message as they may be eligible for our testing discount program.

Otherwise, we like to encourage all of our customers to submit an Abreview via the online product datasheet. We always appreciate customer feedback, whether positive or negative, and we make all product information available to researchers. Plus, each Abreview earns Abpoints that can be used for discounts on future purchases or rewards such as Amazon.com gift certificates.

To find out more about our Abreview system, please see the following link:

https://www.abcam.com/abreviews

I hope this information is helpful. Please do not hesitate to contact me for any further advice or information.

Read More
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (Whole skin)
Specification
Whole skin
Fixative
Formaldehyde
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate
Permeabilization
Yes - 0.1% Tween-20
Blocking step
Serum as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 10% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Oct 11 2012

Question

1) Abcam product code ab 10347
2) Abcam order reference number or product batch number : lot
3) Description of the problem : Unable to see any positive staining of thymine dimers.
4) Sample preparation:
Species Mouse (Balb/c)
Type of sample: Fresh frozen sections, perfusion fixed frozen sections, PFA/formalin fixed paraffin embedded sections, cells in culture, other: formalin fixed paraffin embedded sections
Sample preparation
Positive control: Untreated skin irradiated for 30 min with 8Kj/m2 UVB- Collected 30 min and 24 hour post irradiation
5) Fixation step
Yes/No
If yes: Fixative agent and concentration : 10% buffered formalin
Fixation time: 24 H
Fixation temperature 4C
*also tried unfixed (fresh), and acetone/methanol fixed*
6) Antigen retrieval method : Sodium Citrate Buffer (10mM Sodium Citrate, 0.05% Tween 20, pH 6.0)- 30 min @ 95C (in steamer).
Also tried tris-EDTA Buffer (10mM Tris Base, 1mM EDTA Solution, 0.05% Tween 20, pH 9.0
7) Permeabilization method:
Did you do a permeabilization step (details please) or add permeabilizing agent in any dilution buffers?
Permeabilizing agent and concentration: Tried both triton X-100 (0.05%) and tween 20 (0.1%) in PBS: used in wash buffers
8) Blocking agent (eg BSA, serum…): Tried both goat serum and horse serum at 10%
Concentration: 10%
Blocking time 1 hour, 2 hours, overnight Blocking temperature: 1 hour and 2 hours: RT; overnight: 4C
9) Endogenous peroxidases blocked? Yes
Endogenous biotins blocked? Yes (dako kit)
10) Primary antibody (If more than one was used, describe in “additional notes”) :
Concentration or dilution : Tried, 1:50, 1:100, 1:400, 1:1000, 1:2000, 1:4000
Diluent buffer : Tried 1X TBS (10X TBS= (0.5M Tris Base, 9% NaCl, pH 8.4) ) with 10% serum. Also tried a PBS, 2mM EDTA, 1% BSA diluent.
Incubation time: 1 hour, 2 hours, 3 hours, overnight
11) Secondary antibody:
Species: Used DAKO animal research kit to biotinylate primary antibody prior to application to tissue. Secondary used was peroxidise HRP
Reacts against: biotin
Concentration or dilution : as per kit
Diluent buffer : as per kit
Incubation time : 30 min, 1 hour, 2 hours
Fluorochrome or enzyme conjugate: DAB chromagen/ substrate, as per kit.
12) Washing after primary and secondary antibodies:
Buffer: TBST
Number of washes 3-5x 2-5 min washes (tried both)
13) Detection method: Nikon inverted microscope, light microscopy
14) How many times have you run this staining? 7 times
Do you obtain the same results every time? Yes: no staining at all

Read More
Answer

Thank you for taking time to complete our questionnaire and for contacting us. I am sorry to hear this antibody is not providing satisfactory results.

Having reviewed the protocol details, and after all the optimisation attempts, I am afraid there are no protocol tips I could suggest to obtain better results from this antibody in IHC-P. It may well be this antibody does not work in this application.

The discount code was issued for testing the antibody with mouse samples in ICC. Have you had the opportunity to test the antibody in these conditions?


I hope this information is helpful. Please do not hesitate to contact us for further assistance.

Read More

Answer

I am sending you the new discount codes below, but would suggest if you encounter problems with using the codes for an online order, please give us a call so thatCustomer Service can help you with applying the codes to the order.

DISCOUNT CODE: xxx for ab10347
DISCOUNT CODE: xxx for ab23722
EXPIRATION: xxx

Please let me know if I can help you with anything else.

Read More
Application
Immunohistochemistry (Frozen sections)
Sample
Human Tissue sections (Skin)
Specification
Skin
Fixative
Acetone
Permeabilization
No

Abcam user community

Verified customer

Submitted Jul 06 2012

1-10 of 19 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Sign up