Zikv e protein

Polystyrene 96-well plates were coated overnight at 4 °C with 1 µg/mL of ZIKV E protein (Sinobiological) in a sodium carbonate buffer (pH 9.3). Plates were washed three times in PBS with 0.02% Tween 20 and blocked with non-fat dried milk for one hour at 37 °C with PBS, 2% BSA, and 0.02% Tween 20. Serum from hAd5-ZKV-vaccinated mice was serially diluted in PBS and then incubated at 37 °C. Plates were washed four times with PBS containing 0.02% Tween 20 and incubated with an biotin-labeled goat anti-mouse secondary antibody (Jackson Labs) for one hour. After washing and incubation with horseradish peroxidase (HRP)-conjugated streptavidin (Vector Laboratories, Burlingame, USA), plates were developed with tetramethylbenzidine substrate (Dako). The reaction was stopped with the addition of 2 N H₂SO₄, and emission (450 nm) was read with a microplate reader (Molecular Devices, San Jose, USA).

The microtiter plates (Corning, NY, USA) were coated overnight with 5μg/ml of ZIKV E protein (Sino Biological, Beijing, China). Serum samples were 3-fold serially diluted and E binding antibody was detected by ELISA. Endpoint titers were defined as the highest reciprocal serum dilution giving an absorbance more than 2-fold (in mice) or over 5-fold (in marmosets) background values. Log10 end point titers were reported.