The rescue of phage library was done with M13KO7 helper phage (Stratagene) as mentioned previously (44, 48). The phages were then subjected to five rounds of enrichment by bio-panning as described earlier (43). Briefly, RSC3 core protein was coupled to magnetic beads (MyOne Tosyl activated Dynalbeads, Invitrogen) according to the manufacturer’s instructions, and the antigenic integrity of protein after bead coupling was verified by flow cytometry, using bnAb VRC01. Phages were transferred to an eppendorf tube containing 60 µl RSC3 core protein coated magnetic beads and incubated for 1 h at RT with gentle shaking. The unbound phage was removed by washing 10–15 times with 1× PBS with 0.1% Tween 20. The bound phage was eluted with 0.2 M glycine pH 2.2 for 10 min at RT. The eluted phage were neutralized with 1 M Tris-HCl pH 9.2 and immediately added to TG1/HB2151 cells (OD = 0.5) for infection at 37°C without shaking for 30 min and with shaking at 37°C for 30 min. Cells were spun down and plated on 2XTY agar containing chloramphenicol (30 µg/ml). Finally, the individual colonies were picked and grown in 96 well sterile culture plates (Corning) and a glycerol stock of each colony was stored at −70°C. This procedure was again repeated four times to complete five rounds of biopanning.