A Viral Attachment Assay for Screening of Antiviral Compounds

Begin with a multi-well plate containing a pre-chilled human hepatoma cell monolayer.

Treat test wells with a mix of genetically modified hepatitis C virus carrying the luciferase gene and a target compound; treat control wells with the virus alone.

In the test wells, target molecules interact with viral glycoproteins and cell surface receptors, preventing viral attachment.

However, in control wells, viral glycoproteins interact with cell surface receptors, enabling their attachment. A low temperature inhibits viral entry.

Remove the supernatant, introduce a medium, and re-incubate at the optimum temperature, facilitating viral entry.

Upon entry, the virus releases the genetic material, initiating the synthesis of viral proteins and luciferase enzymes, which are secreted into the medium.

Transfer the luciferase-containing supernatant, and introduce a substrate.

The enzyme-substrate interaction produces light in proportion to luciferase levels.

The lower light intensity in the test wells, compared to the control wells, indicates the antiviral property of the target compound.

For the viral attachment assay, plate 1 x 10⁴ cells per well in a 96-well plate, and incubate overnight for c