Expression of the Lassa virus nucleocapsid protein in insect cells infected with a recombinant baculovirus: application to diagnostic assays for Lassa virus infection.
The coding region of the gene for the nucleocapsid protein of Lassa virus has been inserted into the genome of Autographa californica nuclear polyhedrosis virus (AcNPV) using the transfer vector pAcYM1, so that expression of the foreign DNA is under the control of the promoter of the AcNPV polyhedrin gene. Infection of cultured Spodoptera frugiperda cells with recombinant virus resulted in the synthesis of high levels of a protein that was indistinguishable from the authentic Lassa virus protein by SDS gel electrophoresis and immunoblotting with a variety of specific immune sera and monoclonal antibodies (MAbs). The kinetics of appearance of the protein were comparable to those of polyhedrin production in wild-type AcNPV-infected cells. The recombinant material was antigenic when used in ELISA for Lassa virus-specific antibodies, reacting well with MAbs specific for the nucleocapsid protein and with sera from experimentally infected guinea-pigs. The recombinant ELISA was able to clearly distinguish sera from human cases of Lassa fever against a panel of known negative sera of African origin. Recombinant-infected insect cells were an effective substitute for mammalian cells infected with Lassa virus itself in the immunofluorescence assay for Lassa virus-specific antibodies. This system offers attractive alternatives to the use of Lassa virus-infected materials as reagents in diagnostic procedures.
Public Health Laboratory Service, Centre for Applied Microbiology and Research, Salisbury, U.K.
G N
GN
Barber
J C
JC
Clegg
G
G
Lloyd
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis*; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology; D001116:Arenaviridae / Q000302:isolation & purification*; D001483:Base Sequence; D015139:Blotting, Southern; D002213:Capsid / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D002478:Cells, Cultured; D004279:DNA, Viral / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D015967:Gene Expression Regulation, Viral; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D007304:Insect Viruses / Q000235:genetics; D007835:Lassa Fever / Q000175:diagnosis; D007836:Lassa virus / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000648:ultrastructure; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009036:Moths; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D015183:Restriction Mapping; D014162:Transfection; D014758:Viral Core Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology
D000914:Antibodies, Viral; D000956:Antigens, Viral; D004279:DNA, Viral; D011994:Recombinant Proteins; D014758:Viral Core Proteins