Abstract
Different serological assays were rapidly generated to study humoral responses against the SARS-CoV-2 Spike glycoprotein. Due to the intrinsic difficulty of working with SARS-CoV-2 authentic virus, most serological assays use recombinant forms of the Spike glycoprotein or its receptor binding domain (RBD). Cell-based assays expressing different forms of the Spike, as well as pseudoviral assays, are also widely used. To evaluate whether these assays recapitulate findings generated when the Spike is expressed in its physiological context (at the surface of the infected primary cells), we developed an intracellular staining against the SARS-CoV-2 nucleocapsid (N) to distinguish infected from uninfected cells. Human airway epithelial cells (pAECs) were infected with authentic SARS-CoV-2 D614G or Alpha variants. We observed robust cell-surface expression of the SARS-CoV-2 Spike at the surface of the infected pAECs using the conformational-independent anti-S2 CV3-25 antibody. The infected cells were also readily recognized by plasma from convalescent and vaccinated individuals and correlated with several serological assays. This suggests that the antigenicity of the Spike present at the surface of the infected primary cells is maintained in serological assays involving expression of the native full-length Spike.
Highlights
Very shortly after SARS-CoV-2 was declared a pandemic by the World Health Organization [1], major efforts to understand humoral responses against this new virus were undertaken
Convalescent plasma and plasma from vaccinated individuals were obtained from donors who consented to participating in this research project at CHUM
The plasmid expressing the SARS-CoV-2 Spike glycoprotein was kindly provided by Stefan Pöhlmann (Georg-August University, Göttingen, Germany) and has been previously reported [22]
Summary
Very shortly after SARS-CoV-2 was declared a pandemic by the World Health Organization [1], major efforts to understand humoral responses against this new virus were undertaken. A myriad of different assays were deployed around the globe, including soluble recombinant forms of the Spike, its receptor-binding domain (RBD), cell-based assays expressing different forms of the Spike, pseudoviral assays, and assays using authentic SARS-CoV-2 and infected cells [2,3,4,5,6,7,8,9,10] These assays were used to study vaccine-elicited humoral responses [11,12,13] and to compare these with those elicited by natural infection [11,12,14]. Some of these Fc-mediated effector functions, such as antibody-dependent cellular cytotoxicity (ADCC) or antibody-dependent cellular phagocytosis (ADCP), require recognition of the antigen at the surface of the infected cells This raises an important question about expression levels of SARS-CoV-2 Spike at the surface of primary human airway epithelial cells (pAECs) infected with authentic virus. Measurements were taken with plasma from a cohort of individuals who were SARS-CoV-2 naïve and vaccinated and of those who were previously infected
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