Immunolon 4 hbx microtiter 96 well strips

Immunolon 4 HBX microtiter 96-well strips (VWR) were coated with 150 ng per well of recombinant ZIKV E protein (Cat #:MBS596088; MyBioSource) or recombinant H1 hemagglutinin of A/Puerto Rico/8/1934 (H1N1) (NR-19240, BEI) in bicarbonate/carbonate coating buffer overnight at 4 °C. The plates were blocked with 2% BSA in PBS for 2 hours at room temperature (RT). Sera was serially diluted in 1% BSA in PBS and incubated for 2 hours at RT. The plates were washed 6X with PBST and incubated with goat anti-mouse-HRP antibody (1:5000; Thermo Fisher) in 1% BSA in PBS for 1 hour at RT. After washing 4X with PBST and 2X with PBS, the plate was developed with 1-Step Ultra TMB-ELISA (Thermo Fisher) and the reaction was stopped with 2 M sulfuric acid. The OD450 was detected using a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices) and the endpoint titer was determined as signal that was two and a half times background values.

Immunolon 4 HBX microtiter 96-well strips (VWR) were coated with 150 ng of recombinant ZIKV E protein (Cat #:MBS596088; MyBioSource) in bicarbonate/carbonate coating buffer overnight at 4 °C. The plates were blocked with 2.0% BSA in PBS for 2 hours at room temperature (RT). Sera was diluted in 1.0% BSA in PBS and incubated for 2 hours at RT. The plates were washed 6X with PBST and incubated with goat anti-mouse-HRP antibody (1:2000; Thermo Fisher) in 1.0% BSA in PBS for 1 hour at RT. After washing 4X with PBST and 2X with PBS, the plate was developed with 1-Step Ultra TMB-ELISA (Thermo Fisher), and the reaction was stopped with 2 M sulfuric acid. The OD450 was detected using a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices), and the endpoint titer was determined as signal that was two times background values.