8-Nitroguanosine is a nitrated base of DNA and RNA. It is formed by peroxynitrite, which is generated from nitric oxide and superoxide anion radical. It is known that a large amount of nitric oxide molecules and superoxide anion, generated by inflammation, causes nitration of guanosine. Since chemically modified nucleotides cause mutation during DNA replication, 8-nitroguanosine is thought to be one of the markers of DNA damage related to mutation and cancer. Because of its very high specificity, monoclonal antibody NO2G52 recognizes 8-nitroguanine and 8-nitroguanosine, but it does not cross-react with normal nucleotide bases, 8-hydroxyguanine, 8-hydroxydeoxyguanosine, 3-nitrotyrosine, xanthine, or 2-nitroimidazole (Fig. 1). The specificity of NO2G52 was determined by a competitive ELISA using an 8-nitroguanosine-BSA-coated plate. As shown in the figures below, NO2G52 has very high affinity for 8-nitroguanine and 8-nitroguanosine, and it slightly cross-reacts with 8-bromoguanosine, 8-bromoguanine, and 8-chloroguanine. Anti-Nitroguanosine polyclonal antibody also recognizes 8-nitroguanine and 8-nitroguanosine, but it does not cross-react with normal guanosine, guanine, 8-hydroxyguanine, or 3-nitrotyrosine. Since this antibody was prepared using rabbits, it can be used for immuno-histostaining of rodent tissues such as mice or rats.
Fig 1. Specificity of Anti 8-Nitroguanosine monoclonal antibody
Fig 2 Tissue Staining with Anti-Nitroguanosine Antibody
A) Tissue section (8 days postinfection, influenza) stained with Anti-Nitroguanosine Antibody.
B) The same section viewed using a confocal laser scanning microscope (Fluoroview FV300, Olympus, Nagano, Japan). Strong fluorescence, due to emission of Vector red, is evident in the cytosol.
1) T. Akaike, et al., 8-nitroguanosine formation in viral pneumonia and its implication for pathogenesis, PNAS. 2003;100:685-690.
2) J. Yoshitake, et al., Nitric oxide as an endogenous mutagen for Sendai virus without antiviral activity. J Virol. 2004;78:8709-8719.
3) T. Sawa, et al., Protein S-guanylation by the biological signal 8-nitroguanosine 3 E5 Ecyclic monophosphate. Nat Chem Biol. 2007;3:727-735.
4) M. H. Zaki, et al., Cytoprotective function of heme oxygenase 1 induced by a nitrated cyclic nucleotide formed during murine salmonellosis. J Immunol. 2009;182:3746-3756.
5) Y. Terasaki, et al., Guanine nitration in idiopathic pulmonary fibrosis and its implication for carcinogenesis. Am J Respir Crit Care Med. 2006;174:665-673.
6) T. Sawa, et al., Analysis of urinary 8-nitroguanine, a marker of nitrative nucleic acid damage, by high-performance liquid chromatography-electrochemical detection coupled with immunoaffinity purification: association with cigarette smoking. Free Radic Biol Med. 2006;40:711-720.