Monoclonal Antibodies against DNA Damage [PDF]
DNA damage in cells exposed to ultraviolet (UV) radiation plays significant roles in cell-cycle arrest, activation of DNA repair, cell killing, mutation, and neoplastic transformation. The major types of DNA damage induced by UVB (280-315 nm, component of sunlight) and by UVC (200-280 nm) are cyclobutane pyrimidine dimers (CPDs) and (6-4) photoproducts (6-4PPs), which are formed between adjacent pyrimidine nucleotides on the same strand of DNA. Approximately 70-80% of UV-induced DNA damage is CPDs and the remaining is 6-4PPs and Dewar isomer of 6-4PPs. These types of DNA lesions are repaired by nucleotide excision repair (NER) system in normal human cells. Mori et al (19) have established monoclonal antibodies specific for CPDs or 6-4PPs. These antibodies enable one to quantitate photoproducts in DNA purified from cultured cells or from the skin epidermis using an enzyme-linked immunosorbent assay (ELISA) and to visualize and measure photoproducts in DNA in cultured cells or the skin using indirect immunofluorescence (IIF). This technology would contribute to understanding of molecular mechanisms of cellular responses to UV and DNA damage in many research fields including cancer research, photobiology, dermatology, ophthalmology, immunology, and cosmetology.
The hybridoma was established by fusion of mouse myeloma cells with Balb/c mouse splenocytes immunized with methylated BSA conjugated with calf thymus DNA which was irradiated with UVC. This hybridoma (clone TDM-2) culture supernatant was collected and precipitated with ice-cold ammonium sulfate. After centrifugation, the pellet dissolved in small volume of double-distilled water was dialysed against PBS. The dialysate was then lyophilized.
1) The antibodies bind to CPDs in single-stranded DNA.
2) The antibodies bind to CPDs formed in every dipyrimidine sequence (TT, TC, CT and CC).
3) The antibodies stably binds to CPDs formed in oligonucleotides consisting of more than eight bases.
The antibodies can bind to CPDs in denatured DNA from all organisms from bacteria to human.
Immunocytochemistry; 1:1500 PROTOCOLS< Immunofluorescence microscopy >
ELISA; 1:1000 PROTOCOLS<ELISA>
Western blotting; Not tested
Immunoprecipitation; Not tested
Immunohistochemistry; Not tested
Flow cytometry; Not tested
Induction and repair of UV-induced DNA damage in normal human cells measured by ELISA
Visualization of UV-induced DNA damage by using immunofluorescence
Cosmo Bio's new Cyclobutane Pyrimidine Dimer (CPD) ELISA Kit uses the highly sensitive and specific monoclonal anti-CPD antibody clone TDM-2, established by Mori et al. TDM-2 is widely cited in the literature and is considered to be the gold standard antibody for CPD detection and quantification.Cosmo Bio's CPD ELISA Kit is the first and only commercially available ELISA kit using TDM-2. All kits components are optimized for high sensitivity CPD detection from cultured cells or skin epidermis.
High Sensitivity CPD ELISA kit Ver.2
To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.