Omicron ba 1

We diluted the antigens for antibody detection in PBS and coated the plates (Nunc MaxiSorp, Thermofisher Scientific) with 250 ng/mL WT (AcroBiosystems) or Omicron BA.1 (AcroBiosystems) SARS-CoV-2 S protein for IgG and IgG avidity assessment and 500 ng/mL ancestral (Sinobiological) or Omicron BA.1 (AcroBiosystems) S for FcγRIIIa-binding detection. ORF8 protein of 300 ng/mL was coated at 37 for 2 hours. The plates were blocked with 1% FBS in PBS for 1 hour, followed by incubation with 1:100 HI sera diluted in 0.05% Tween-20/0.1% FBS in PBS for 2 hours for IgG detection, and 1:50 for 1 hour at 37 for FcγRIIIa-binding detection, prior to rinsing. For avidity, plates with 8M were washed with urea 3 times. IgG was measured after 2 hours of incubation period with anti-IgG-HRP (1:5000; G18-145, BD), HRP revealed with addition of stabilized hydrogen peroxide and tetramethylbenzidine (R&D systems) for 20 minutes. The reaction was terminated with 2N H2SO4, which was then analyzed at 450 nm wavelength with an absorbance microplate reader (Tecan Life Sciences). Similarly, FcγRIIIa-binding antibodies were assessed after incubation with biotinylated FcγRIIIa-V158 at 100 ng/mL for 1 hour at 37 after streptavidin-HRP (1:10000, Pierce).