Intracellular interference of tick-borne flavivirus infection by using a single-chain antibody fragment delivered by recombinant Sindbis virus.
A single-chain antibody fragment that identifies a neutralizing epitope on the envelope protein of louping ill and some other tick-borne flaviviruses was previously expressed in soluble form from bacteria and shown to be functionally active in vitro. To see whether or not the single-chain antibody could bind and inactivate infectious virus in vivo, we have used recombinant Sindbis virus as a delivery vehicle for intracellular expression of the antibody fragment. The variable genes and interchain linker encoding the single-chain antibody were cloned into a double subgenomic Sindbis virus expression vector to generate recombinant Sindbis virus. Infection with this recombinant Sindbis virus provided high-level cytoplasmic expression of the antibody fragment in mammalian cells. We demonstrate (i) that the antibody fragment was antigen binding and (ii) that louping ill virus infectivity was significantly reduced in the presence of intracellular antibody expressed by the superinfecting recombinant Sindbis virus.
Jiang
W
W
Natural Environment Research Council Institute of Virology and Environmental Microbiology, Oxford, United Kingdom.
Venugopal
K
K
Gould
E A
EA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D004669:Encephalitis Viruses, Tick-Borne / Q000276:immunology*; D007128:Immunoglobulin Fragments / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000276:immunology*; D012845:Sindbis Virus / Q000235:genetics*; D013552:Swine; D014762:Viral Interference; D014764:Viral Proteins / Q000378:metabolism; D014779:Virus Replication
D007128:Immunoglobulin Fragments; D011994:Recombinant Proteins; D014764:Viral Proteins; C055966:immunoglobulin Fv