Screening and Identifying a Novel ssDNA Aptamer against Alpha-fetoprotein Using CE-SELEX.
Alpha-fetoprotein (AFP) is a liver cancer associated protein and has long been utilized as a serum tumor biomarker of disease progression. AFP is usually detected in HCC patients by an antibody based system. Recently, however, aptamers generated from systematic evolution of ligands by exponential enrichment (SELEX) were reported to have an alternative potential in targeted imaging, diagnosis and therapy. In this study, AFP-bound ssDNA aptamers were screened and identified using capillary electrophoresis (CE) SELEX technology. After cloning, sequencing and motif analysis, we successfully confirmed an aptamer, named AP273, specifically targeting AFP. The aptamer could be used as a probe in AFP immunofluorescence imaging in HepG2, one AFP positive cancer cell line, but not in A549, an AFP negative cancer cell line. More interesting, the aptamer efficiently inhibited the migration and invasion of HCC cells after in vivo transfection. Motif analysis revealed that AP273 had several stable secondary motifs in its structure. Our results indicate that CE-SELEX technology is an efficient method to screen specific protein-bound ssDNA, and AP273 could be used as an agent in AFP-based staining, diagnosis and therapy, although more works are still needed.
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.
Lili
L
Dong
Department of Laboratory Medicine, Zhongshan Hospital, Fudan University, Shanghai 200032, China.
Qiwen
Q
Tan
School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.
Wei
W
Ye
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.
Dongli
D
Liu
School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.
Haifeng
H
Chen
Shanghai Aijin Biochemical Science &Technology Co. Ltd., Shanghai 200336, China.
Hongwei
H
Hu
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.
Duo
D
Wen
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.
Yang
Y
Liu
Cancer Research Institute, Xiangya School of Medicine, Central South University, Key Laboratory of Chinese Ministry of Education, Changsha 410078, China.
Ya
Y
Cao
Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032, China.
Jingwu
J
Kang
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.|Institute of Biomedical Sciences, Fudan University, Shanghai 200032, China.
Jia
J
Fan
Department of Laboratory Medicine, Zhongshan Hospital, Fudan University, Shanghai 200032, China.
Wei
W
Guo
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Key Laboratory of Carcinogenesis and Cancer Invasion, Ministry of Education, Shanghai 200032, China.
Weizhong
W
Wu
D052157:Aptamers, Nucleotide / Q000737:chemistry*; D014408:Biomarkers, Tumor / Q000032:analysis*; D045744:Cell Line, Tumor; D004277:DNA, Single-Stranded / Q000737:chemistry; D019075:Electrophoresis, Capillary; D006801:Humans; D009369:Neoplasms / Q000175:diagnosis; D052156:SELEX Aptamer Technique; D000509:alpha-Fetoproteins / Q000032:analysis* / Q000037:antagonists & inhibitors
D052157:Aptamers, Nucleotide; D014408:Biomarkers, Tumor; D004277:DNA, Single-Stranded; D000509:alpha-Fetoproteins