Meliana Eka Saputri et al.

Coronavirus disease (COVID)-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has become a global pandemic disease that has social and economic chaos. An alternative mitigation strategy may involve the use of specific immunoglobulin (Ig)-Y derived from chicken eggs. Our study aimed to evaluate the neutralizing potential of specific IgY targeting S1, receptor-binding-domain (RBD) of spike glyco-protein and nucleocapsid (N) of SARS-CoV-2 to inhibit RBD and angiotensin-converting-enzyme-2 (ACE2) binding interaction. Hy-Line Brown laying hens were immunized with recombinant S1, RBD spike glycoprotein, and nucleocapsid (N) of SARS-CoV-2. The presence of specific S1,RBD,N-IgY in serum and egg yolk was verified by indirect enzyme-linked immunosorbent assay (ELISA). Specific S1,RBD,N-IgY was purified and characterized from egg yolk using sodium-dodecyl-sulfate-polyacrylamide-gel-electrophoresis (SDS-PAGE), and was subsequently evaluated for inhibition of the RBD-ACE2 binding interaction in vitro. Specific IgY was present in serum at 1 week post–initial immunization (p.i.i), whereas its present in egg yolk was confirmed at 4 weeks p.i.i. Specific S1,RBD,N-IgY in serum was able to inhibit RBD-ACE2 binding interaction between 4 and 15 weeks p.i.i. The results of the SDS-PAGE revealed the presence of bands with molecular weights of 180 kDa, indicating the presence of whole IgY. Our results demonstrated that S1,RBD,N-IgY was able to inhibit RBD-ACE2 binding interaction in vitro, suggesting its potential use in blocking virus entry. Our study also demonstrated proof-of- concept that laying hens were able to produce this speci c IgY, which could block the viral binding and large production of this speci c IgY is feasible.