Several HIV cell fusion assays have been established previously. In term of
components used to generate the fusion assay, there are two fusion systems. One is a viruscell
system that uses viral particles or recombinant viruses to mediate cell fusion. The other is
a cell-cell system that uses two cell types expressing HIV-1 Env (effector cells) and human
CD4 and co-receptors (target cells). The cell-cell fusion systems are more preferable than the
virus-cell system because no infectious viruses are produced. As a result of successful cell
fusion, multinucleated cells or syncytia are formed. Therefore, the extent of fusion could be
determined by the amount of syncytial formations. A conventional method to quantify fusion
is to directly count number of syncytial cells; however, it is laborious and time-consuming.
An improved approach to assess fusion events is to apply reporter proteins such as firefly
2
luciferase, β-galactosidase, and green fluorescent protein (GFP). These reporter genes are
specifically controlled by either bacteriophage T7 promoter or HIV-1 long terminal repeat
(LTR) promoter. For the reporter gene driven by T7 promoter, the gene would be expressed
only after acquiring T7 RNA polymerase whereas those driven by HIV-1 LTR, the gene
expression would be greatly enhanced after gaining trans-activator of transcription (Tat)
protein from the fused cell partner.5-13 The enzyme-based systems, however, require
additional steps of cell lysate preparation and enzyme substrate addition prior to obtaining a
fusion readout.
Several HIV cell fusion assays have been established previously. In term ofcomponents used to generate the fusion assay, there are two fusion systems. One is a viruscellsystem that uses viral particles or recombinant viruses to mediate cell fusion. The other isa cell-cell system that uses two cell types expressing HIV-1 Env (effector cells) and humanCD4 and co-receptors (target cells). The cell-cell fusion systems are more preferable than thevirus-cell system because no infectious viruses are produced. As a result of successful cellfusion, multinucleated cells or syncytia are formed. Therefore, the extent of fusion could bedetermined by the amount of syncytial formations. A conventional method to quantify fusionis to directly count number of syncytial cells; however, it is laborious and time-consuming.An improved approach to assess fusion events is to apply reporter proteins such as firefly2luciferase, β-galactosidase, and green fluorescent protein (GFP). These reporter genes arespecifically controlled by either bacteriophage T7 promoter or HIV-1 long terminal repeat(LTR) promoter. For the reporter gene driven by T7 promoter, the gene would be expressedonly after acquiring T7 RNA polymerase whereas those driven by HIV-1 LTR, the geneexpression would be greatly enhanced after gaining trans-activator of transcription (Tat)protein from the fused cell partner.5-13 The enzyme-based systems, however, requireadditional steps of cell lysate preparation and enzyme substrate addition prior to obtaining a
fusion readout.
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