UV-induced DNA damage monoclonal antibodies

Powerful Monoclonal Antibodies to

UV-induced DNA Damage

Anti Cyclobutane Pyrimidine Dimers (CPDs), clone TDM-2
Anti (6-4) Photoproducts (6-4 PPs), clone 64M-2
Anti Dewar Photoproducts (Dewar PPs), clone DEM-1

Quantitate photoproducts by ELISA or visualize by immunofluorescence.
Specific for nominal UV-induced DNA lesion.
Detect photoproducts in single stranded DNA or oligos (8bp or greater).
Reacts with photoproducts from any species.
Widely cited.

Background

Prolonged exposure to solar UV radiation may result in acute and chronic health effects on the skin, eye, and immune system, including skin cancers. These harmful effects are suggested to deeply relate to DNA damage.

The major types of DNA damage induced by solar UV radiation are cyclobutane pyrimidine dimers (CPDs), (6–4) photoproducts (6-4PPs), and Dewar photoproducts (DewarPPs), which are formed between adjacent pyrimidine nucleotides on the same strand of DNA. These helix-distorting DNA lesions are repaired exclusively by a nucleotide excision repair system in humans.

Mori et. al. have developed and characterized monoclonal antibodies specific for CPDs or 6-4PPs (1) while Matsunaga et. al. have established and characterized monoclonal antibodies against DewarPPs (2). These antibodies enable quantitation of photoproducts in DNA purified from cultured cells or from skin epidermis using an enzyme-linked immunosorbent assay (ELISA), and to visualize and measure photoproducts in DNA of cultured cells or skin by indirect immunofluorescence. These capabilities will contribute to understanding the molecular mechanism of cellular responses to UV and DNA damage in many research fields including cancer research, photobiology, dermatology, ophthalmology, immunology, and cosmetology.

(1) Toshio Mori, Misa Nakane, Tsuyoshi Hattori, Tsukasa Matsunaga, Makoto Ihara, Osamu Nikaido. Simultaneous establishment of monoclonal antibodies specific for either cyclobutane pyrimidine dimer or (6-4) photoproduct from the same mouse immunized with ultraviolet-irradiated DNA. Photochem. Photobiol., 54: 225-232 (1991). PMID 1780359

(2) Tsukasa Matsunaga, Yuri Hatakeyama, Michi Ohta, Toshio Mori and Osamu Nikaido. Establishment and characterization of a monoclonal antibody recognizing the Dewar isomers of (6-4) photoproducts. Photochem. Photobiol., 57: 934-940 (1993). PMID 8367534

Applications

Immunohistochemistry

The technique of micropore UV irradiation combined with fluorescent antibody labeling is very powerful for examining whether a protein of interest is recruited to the sites of UV-induced DNA damage. Micropore UV irradiation induces UV-damage at localized areas of nuclei using a polycarbonate isopore membrane filter. The polycarbonate blocks UV radiation, and cells are exposed only through the 5 µm pores of the filter. 0.5 h after micropore UV irradiation, cells were fixed and Immunofluorescent double staining for DNA damage and repair protein were performed.

Cells were doubly stained for XPB and for CPD 0.5 h after local UV irradiation. In normal MSU-1 cells, XPB foci overlapped with the corresponding CPD foci, indicating that XPB is quickly recruited to the sites of DNA damage for repair. In contrast, no or less bright XPB foci at the DNA damage sites were observed in repair deficient TTD cell lines.

ELISA

Genomic DNA is purified from UV-damaged cells and denatured DNA is coated on wells of 96-well plate. The binding of TDM-2 or 64M-2 to DNA damage is detected by sequential treatment with biotinylated 2nd antibody and streptavidin-peroxidase. Then, the absorbance of colored products derived from OPD is measured at 492 nm.

Normal human cells repair 90% of the initial 6-4PP within 3 h after UV irradiation, while they remove 50% of the initial CPD within 24 h. Both damage are repaired by the same nucleotide excision repair (NER) pathway, but 6-4PP forms bigger distortion in DNA than CPD does, resulting in much more efficient repair. In contrast, repair deficient XP-C cells can not repair both damage at all.

Products

Product	Host	Clone	Application	Catalog No/ Price
Anti CPDs	Mouse	TDM-2	ELISA, IF	CAC-NM-DND-001 $550
Anti 6-4 PPs	Mouse	64M-2	ELISA, IF	CAC-NM-DND-002 $550
Anti Dewar PPs	Mouse	DEM-1	ELISA, IF	CAC-NM-DND-003 $550