title: Development of an enzyme-linked immunospot assay for determination of rotavirus infectivity.
abstract: Conventional rotavirus infectivity assays are time consuming, labor intensive, and with low sample throughput. To overcome these problems, a 96-well microplate enzyme-linked immunospot assay (Elispot) was developed for the measurement of rotavirus infectious titers. The infected MA104 cells were stained with a horseradish peroxidase-conjugated anti-VP6 monoclonal antibody followed by detection with an ELISPOT analyzer. A linear relationship was found between spot number and input of rotavirus dose in SA11 and 10 rotavirus isolates of different genotypes. The propagation of rotavirus SA11 in MA104 cells was monitored, and the neutralizing activity of serum samples and monoclonal antibodies was determined. The 50% neutralizing titer (NT50) of serum and 50% inhibitory concentration (IC50) of monoclonal antibodies were correlated well with the results determined by ELISA-based neutralization assay. In conclusion, a rapid and semi-automated procedure to determine rotavirus infectivity was developed, which will be useful to study the infectivity and the neutralizing epitopes of rotavirus.
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China.
authors.forename: Tingdong
authors.initials: T
authors.lastname: Li
authors.affiliation: School of Life Science, Xiamen University, Xiamen, China.
authors.forename: Haijun
authors.initials: H
authors.lastname: Lin
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China.
authors.forename: Linqi
authors.initials: L
authors.lastname: Yu
authors.affiliation: School of Life Science, Xiamen University, Xiamen, China.
authors.forename: Miaoge
authors.initials: M
authors.lastname: Xue
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China. Electronic address: sxge@xmu.edu.cn.
authors.forename: Shengxiang
authors.initials: S
authors.lastname: Ge
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China.
authors.forename: Qinjian
authors.initials: Q
authors.lastname: Zhao
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, National Institute of Diagnostics and Vaccine Development in Infectious Disease, School of Public Health, Xiamen University, Xiamen, China; School of Life Science, Xiamen University, Xiamen, China.
authors.forename: Ningshao
authors.initials: N
authors.lastname: Xia
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood / Q000276:immunology; D002460:Cell Line; D002522:Chlorocebus aethiops; D058501:Enzyme-Linked Immunospot Assay / Q000379:methods*; D006735:Horseradish Peroxidase / Q000032:analysis; D020128:Inhibitory Concentration 50; D009500:Neutralization Tests; D012401:Rotavirus / Q000276:immunology / Q000302:isolation & purification*; D013194:Staining and Labeling / Q000379:methods; D019562:Viral Load
chemical_list: D000914:Antibodies, Viral; D006735:Horseradish Peroxidase
keywords: Elispot; Infectivity; Neutralization; Rotavirus