Effective therapies are urgently needed for the SARS-CoV-2/COVID19 pandemic. We identified panels of fully human monoclonal antibodies (mAbs) from eight large phagedisplayed Fab, scFv and VH libraries by panning against the receptor binding domain (RBD) of the SARS-CoV-2 spike (S) glycoprotein. One high affinity mAb, IgG1 ab1, specifically neutralized live SARS-CoV-2 with exceptional potency as measured by two different assays. It competed with human angiotensin-converting enzyme 2 (hACE2) for binding to RBD suggesting a competitive mechanism of virus neutralization. IgG1 ab1 protected transgenic mice expressing hACE2 from high-titer intranasal SARS-CoV-2 challenge (10 5 plaque forming units). Another antibody, VH ab5 did not compete with hACE2 and ab1, and did not neutralize SARS-CoV-2 although its affinity was comparable to that of ab1. The ab1 sequence has relatively low number of somatic mutations indicating that ab1-like antibodies could be quickly elicited during natural SARS-CoV-2 infection or by RBD-based vaccines. IgG1 ab1 does not have developability liabilities, and thus has potential for therapy and prophylaxis of SARS-CoV-2 infections. The rapid identification (within 6 days) of potent mAbs shows the value of large antibody libraries for response to public health threats from emerging microbes.The severe acute respiratory distress coronavirus 2 (SARS-CoV-2) (1) has spread worldwide thus requiring safe and effective prevention and therapy. Inactivated serum from convalescent patients inhibited SARS-CoV-2 replication and decreased symptom severity of newly infected patients (2, 3) suggesting that monoclonal antibodies (mAbs) could be even more effective.Human mAbs are typically highly target-specific and relatively non-toxic. By using phage display we have previously identified a number of potent fully human mAbs (m396, m336, m102.4)
Control of small molecule hapten epitope densities on antigenic carrier proteins is essential for development and testing of optimal conditions for vaccines. Yet, accurate determination of epitope density can be extremely difficult to accomplish, especially with the use of small haptens, large molecular weight carrier proteins, and limited amounts of protein. Here we report a simple radiometric method that uses 14C-labeled cystine to measure hapten epitope densities during sulfhydryl conjugation of haptens to maleimide activated carrier proteins. The method was developed using a (+)-methamphetamine (METH)-like hapten with a sulfhydryl terminus, and two prototype maleimide activated carrier proteins, bovine serum albumin (BSA) and immunocyanin monomers of keyhole limpet hemocyanin. The method was validated by immunochemical analysis of the hapten–BSA conjugates, and least-squares linear regression analysis of epitope density values determined by the new radiometric method versus values determined by matrix-assisted laser desorption/ionization mass spectrometry. Results showed that radiometric epitope density values correlated extremely well with the mass spectrometrically derived values (r2 = 0.98, y = 0.98x + 0.91). This convenient and simple method could be useful during several stages of vaccine development including the optimization and monitoring of conditions for hapten–protein conjugations, and choosing the most effective epitope densities for conjugate vaccines.
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