title: Probing cigarette smoke-induced DNA single-strand breaks and screening natural protective compounds by use of magnetic bead-based chemiluminescence.
abstract: Magnetic bead (MB)-based chemiluminescence (CL) ELISA can be a sample-thrifty, time-saving tool for evaluation of cigarette smoke-induced DNA single-strand breaks (SSBs) with high specificity. This article describes a novel approach using immobilized oligonucleotide on MBs to determine cigarette smoke-induced DNA SSBs and screen some protective natural compounds. Typically, fluorescein-labeled DNA (FAM-DNA) was immobilized on the MBs and then oxidized by the smoke in the absence or presence of natural compounds, and a part of FAM-DNA was fragmented due to cigarette smoke-induced DNA SSB and then detached from MBs whereas other non-broken FAM-DNA still remained on MBs. Then, any broken FAM-DNA fragments, complex tobacco smoke matrix, and other stuff related with natural compounds were conveniently washed away by a magnetic force, and thus possible interfering substances were completely removed. Finally, those remaining non-broken FAM-DNA on MBs were reacted with HRP-labeled anti-fluorescein antibody and then detected by CL ELISA. CL signal was converted to molar concentrations of the FAM-DNA by interpolation from a pre-determined standard linear calibration curve. The level of DNA SSBs induced by cigarette smoke was thus calculated using the method. A library of 30 natural products was subsequently screened, and two among them were found to protect DNA from oxidative damage and thus may be promising compounds for the development of new drugs. The method developed will be useful for quantitative screening of drug genotoxicity in terms of induction of DNA SSBs. Graphical abstract ᅟ.
authors.affiliation: Yangpu Hospital, Tongji University School of Medicine, No. 1239, Siping Road, 200090, Shanghai, China.
authors.forename: Hongjun
authors.initials: H
authors.lastname: Chen
authors.affiliation: Yangpu Hospital, Tongji University School of Medicine, No. 1239, Siping Road, 200090, Shanghai, China.
authors.forename: Zicheng
authors.initials: Z
authors.lastname: Yu
authors.affiliation: School of Pharmacy, Fudan University, No. 826 Zhangheng Road, 200032, Shanghai, China.
authors.forename: Zhijuan
authors.initials: Z
authors.lastname: Cao
authors.affiliation: School of Pharmacy, Fudan University, No. 826 Zhangheng Road, 200032, Shanghai, China. cyliu@shmu.edu.cn.
authors.forename: Choiwan
authors.initials: C
authors.lastname: Lau
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001688:Biological Products / Q000494:pharmacology*; D045744:Cell Line, Tumor; D000073865:Cigarette Smoking / Q000009:adverse effects*; D053904:DNA Breaks, Single-Stranded / Q000187:drug effects*; D055808:Drug Discovery / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004847:Epithelial Cells / Q000187:drug effects; D008163:Luminescent Measurements / Q000379:methods*; D008280:Magnetics; D051379:Mice; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics; D010957:Plasmids; D012906:Smoke / Q000009:adverse effects* / Q000032:analysis
chemical_list: D000975:Antioxidants; D001688:Biological Products; D012906:Smoke
keywords: Chemiluminescence; DNA; Magnetic beads; Natural products; Screening