Expression and characterization of human group C rotavirus virus-like particles in insect cells.
Group C rotavirus (GpC RV) is a causative agent of acute gastroenteritis in children and adults. We expressed the three major capsid proteins VP2, VP6 and VP7 of human GpC RV in baculovirus and demonstrated the self-assembly of VP2/6/7 or VP6/7 virus-like particles (VLPs) in insect cells. We examined a number of parameters, including the kinetics of protein synthesis in different cell lines and media, to optimize the most favorable conditions for the synthesis of recombinant viral proteins and the production of VLPs in Sf9 cells. Hyperimmune serum to VP2/6/7 and VP6/7 VLPs recognized individual recombinant proteins of human GpC RV by Western blot analysis. This serum also showed specific reactivities with the corresponding GpC VLPs but not GpA RV by using immune electron microscopy (IEM) and enzyme immunoassay (EIA). The ability to produce an unlimited amount of GpC RV antigen and the availability of high quality antibody will allow us to develop sensitive and specific diagnostic assays to better determine the epidemiology and disease burden of GpC RV in humans.
Clark
Kristina B
KB
National Center for Immunization and Respiratory Diseases, Atlanta, Georgia, USA.
Lin
Seh-Ching
SC
Humphrey
Charles
C
Foytich
Kimberly
K
Esona
Mathew
M
Wang
Yuhuan
Y
Liu
Merry
M
Jiang
Baoming
B
D000818:Animals; D000914:Antibodies, Viral / Q000096:biosynthesis / Q000276:immunology; D000918:Antibody Specificity; D000956:Antigens, Viral / Q000032:analysis / Q000096:biosynthesis* / Q000276:immunology; D016367:Baculoviridae / Q000235:genetics; D015153:Blotting, Western; D036022:Capsid Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000276:immunology; D002460:Cell Line; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007313:Insecta / Q000821:virology; D016253:Microscopy, Immunoelectron; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis / Q000276:immunology; D012401:Rotavirus / Q000276:immunology* / Q000502:physiology / Q000648:ultrastructure; D014771:Virion / Q000276:immunology / Q000502:physiology / Q000648:ultrastructure; D019065:Virus Assembly
D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; D007106:Immune Sera; D011994:Recombinant Proteins; C442545:VP2 protein, Rotavirus; C061332:VP6 protein, Rotavirus; C050779:VP7 protein, Rotavirus