Extended Data Fig. 3. Functional assays from single antigen-reactive B cells.
a. Schematic of detection of antigen-specific antibody. Biotinylated antigen (dark grey) was coupled to a streptavidin-conjugated polystyrene bead (light grey). Antibodies (blue) are secreted by single B cells loaded into individual NanoPens on the Berkeley Lights Beacon optofluidic device. Antibody binding to antigen was detected with a fluorescent anti-human IgG secondary Ab (black).
b. Left: Schematic of fluorescing beads in the channel above a pen containing an individual B cell indicates antigen-specific reactivity.
Top right: False-color still image of positive wells with B cells secreting S2Pecto-reactive antibodies. Reactive antibody diffusing out of a pen is visualized as a plume of fluorescence.
Bottom right: False-color still image of positive wells with B cells secreting RBD-mFc-reactive antibodies.
c. Representative images of RBD-mFc reactive B cells from a single-B-cell secretion assay
d. Identification of mAbs with hACE2-blocking activity using single-cell functional screening. Left: Schematic illustrating detection of secreted Ab and hACE2 binding on an RBD-mFc-coated streptavidin bead. Ab binding was detected in one fluorescent channel, while hACE2 binding was detected in another fluorescent channel. The top panel illustrates an RBD-binding, non-blocking mAb, where the bead is positive for both Ab and hACE2 signals, while the bottom panel illustrates an RBD-binding mAb that competes with hACE2 for binding, where the bead is positive for only Ab signal. Right: Representative images of a B cell secreting non-blocking Abs (top) and a B cell secreting hACE2-blocking mAbs (bottom). Streptavidin beads are loaded into the same pens as B cells. The fluorescence of the streptavidin beads in the same pen as the B cell secreting hACE2-blocking Abs is reduced relative to adjacent wells, indicating hACE2-blocking activity.