Figure 6.
The multiplexed assay can be expanded to primary neutrophils. Crimson, blue, and yellow-green 1 μm fluorescent beads were coupled to HIV gp120, influenza HA, and Ebola glycoprotein antigen, respectively. Three plasma samples from HIV seronegative and positive donors were titrated from 1:25 to 1:1,600 and tested in the phagocytosis assay using each antigen-coupled bead set individually and in the 3-bead multiplexed format. Instead of THP-1 cells, primary leukocytes were used as a source of neutrophils to measure antibody-dependent phagocytosis. (A,C) Titration curves show phagocytic scores for gp120- (A) and HA- (C) specific ADNP in the multiplexed and single-bead formats for representative HIV-positive and HIV-negative plasma samples. Each point represents the mean ± SD of triplicate wells. The dashed line indicates the phagocytic score for a no-antibody control. (B,D) Dot plots show phagocytic scores for gp120- (B) and HA- (D) specific ADNP using 1 vs. 3 bead-sets in a single well. R2 values were computed with linear regression Pearson correlation with a 95% confidence interval.