Detection of B virus antibody in monkey sera using glycoprotein D expressed in mammalian cells.
The gene encoding glycoprotein D (gD) of the monkey B virus (Cercopithecine herpesvirus 1) was cloned into a mammalian expression vector, pcDNA3.1(-), and the recombinant plasmid DNA was transfected into COS7 cells. The expression of gD in transfected COS7 cells was detected by indirect immunofluorescence assay or radioimmunoprecipitation analysis (RIPA). Although the expressed gD protein was revealed to react well with sera from monkeys naturally infected with B virus by RIPA, some sera showed reduced reactivity when analyzed by the Western blotting (WB) method. Some sera also showed relatively high background when the WB was performed using gD expressed from recombinant plasmid. The mutant gD protein lacking the transmembrane domain (TM) and cytoplasmic tail (CT) was next expressed in COS7 cells. The mutant protein was secreted into culture medium without apparent loss of the antigenicity. Using the secretory form of the gD protein as antigen in dot blot analysis, sera from B virus-infected monkeys were shown to react with the mutant protein without nonspecific reaction. Since the recombinant gD or its derivative lacking TM and CT could be expressed in mammalian cells with proper antigenicity, these antigens appeared to be useful for serological detection of B virus infection in monkeys.
Tanabayashi
K
K
Tsukuba Primate Center for Medical Science, National Institute of Infectious Diseases, 1 Hachimandai, Tsukuba, Ibaraki 208-0843, Japan. ktana@nih.go.jp
Mukai
R
R
Yamada
A
A
D000818:Animals; D000914:Antibodies, Viral / Q000097:blood*; D019556:COS Cells; D002522:Chlorocebus aethiops; D017353:Gene Deletion; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D006568:Herpesvirus 1, Cercopithecine / Q000276:immunology*; D008252:Macaca fascicularis; D008992:Monkey Diseases / Q000175:diagnosis* / Q000276:immunology / Q000821:virology; D014162:Transfection; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
D000914:Antibodies, Viral; D014759:Viral Envelope Proteins; C039447:glycoprotein D, Human herpesvirus 1