An In Vitro Assay to Examine the Antibody-Dependent Enhancement of Viral Infection

Take a microplate with serial dilutions of a serum sample containing antibodies against the Zika virus.

Add Dengue reporter virus particles or RVPs, and incubate.

The engineered RVPs comprise an envelope with structural proteins and a nucleocapsid containing genomic  RNA, encoding a fluorescent protein.

The structural homology of Dengue and Zika viruses' envelope proteins causes the antibodies to cross-react with the RVPs, forming a complex.

Add human leukemia cells impermissible to infection by RVPs.

The cells possess Fc receptors that recognize the RVP-bound antibodies, facilitating RVPs' entry inside endosomes — termed antibody-dependent enhancement or ADE.

Pellet the cells and discard the supernatant containing non-internalized RVPs. Add a growth media and incubate.

The pH-dependent fusion of RVP and endosomal membranes releases the nucleocapsid, which disassembles to release genomic RNA, enabling fluorescent protein expression.

Fix the cells and analyze them using flow cytometry.

A higher fluorescence at a suitable antibody concentration represents increased RVP internalization, indicating enhanced infection.