CS-SELEX generates high-affinity ssDNA aptamers as molecular probes for hepatitis C virus envelope glycoprotein E2.
Currently, the development of effective diagnostic reagents as well as treatments against Hepatitis C virus (HCV) remains a high priority. In this study, we have described the development of an alive cell surface--Systematic Evolution of Ligands by Exponential Enrichment (CS-SELEX) technique and screened the functional ssDNA aptamers that specifically bound to HCV envelope surface glycoprotein E2. Through 13 rounds of selection, the CS-SELEX generated high-affinity ssDNA aptamers, and the selected ssDNA aptamer ZE2 demonstrated the highest specificity and affinity to E2-positive cells. HCV particles could be specifically captured and diagnosed using the aptamer ZE2. A good correlation was observed in HCV patients between HCV E2 antigen-aptamer assay and assays for HCV RNA quantities or HCV antibody detection. Moreover, the selected aptamers, especially ZE2, could competitively inhibit E2 protein binding to CD81, an important HCV receptor, and significantly block HCV cell culture (HCVcc) infection of human hepatocytes (Huh7.5.1) in vitro. Our data demonstrate that the newly selected ssDNA aptamers, especially aptamer ZE2, hold great promise for developing new molecular probes, as an early diagnostic reagent for HCV surface antigen, or a therapeutic drug specifically for HCV.
Chen
Fang
F
State Key Laboratory of Virology, Department of Immunology, Wuhan University School of Medicine, Wuhan, People's Republic of China.
Hu
Yilan
Y
Li
Dongqing
D
Chen
Haidan
H
Zhang
Xiao-Lian
XL
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D005838:Genotype; D016174:Hepacivirus / Q000235:genetics; D006526:Hepatitis C / Q000517:prevention & control / Q000821:virology; D022781:Hepatocytes / Q000378:metabolism / Q000821:virology; D006801:Humans; D007700:Kinetics; D051379:Mice; D008856:Microscopy, Fluorescence; D015335:Molecular Probes / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D011485:Protein Binding; D012367:RNA, Viral / Q000378:metabolism; D052156:SELEX Aptamer Technique / Q000379:methods*; D060225:Tetraspanin 28; D014759:Viral Envelope Proteins / Q000378:metabolism*; D014771:Virion / Q000378:metabolism
D015703:Antigens, CD; D052157:Aptamers, Nucleotide; C554798:CD81 protein, human; C554799:Cd81 protein, mouse; D015335:Molecular Probes; D012367:RNA, Viral; D060225:Tetraspanin 28; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus; D016650:Fluorescein-5-isothiocyanate