In order to understand the kinetics, timing and persistence of SARS-CoV2 neutralizing antibodies (Nabs) we used a surrogate viral neutralization test to evaluate their levels in patients with varying severity of illness, in those with prolonged shedding and those with mild/asymptomatic illness at various time points. Patients with severe or moderate COVID-19 illness had earlier appearance of Nabs at higher levels compared to those with mild or asymptomatic illness. Furthermore, those who had prolonged shedding of the virus, had Nabs appearing faster and at higher levels than those who cleared the virus earlier. Although all individuals appeared to be antibody positive by end of week 5, the positivity rates declined thereafter, especially in those who had mild or asymptomatic illness.

The molecular mechanisms regulating antigen translocation into the cytosol for cross-presentation are under controversial debate, mainly because direct data is lacking. Here, we have provided direct evidence that the activity of the endoplasmic reticulum (ER) translocon protein Sec61 is essential for endosome-to-cytosol translocation. We generated a Sec61-specific intrabody, a crucial tool that trapped Sec61 in the ER and prevented its recruitment into endosomes without influencing Sec61 activity and antigen presentation in the ER. Expression of this ER intrabody inhibited antigen translocation and cross-presentation, demonstrating that endosomal Sec61 indeed mediates antigen transport across endosomal membranes. Moreover, we showed that the recruitment of Sec61 toward endosomes, and hence antigen translocation and cross-presentation, is dependent on dendritic cell activation by Toll-like receptor (TLR) ligands. These data shed light on a long-lasting question regarding antigen cross-presentation and point out a role of the ER-associated degradation machinery in compartments distinct from the ER.

We discovered that values for 2 out of 79 antibodies have unfortunately been wrongly reported. For antibody FnC1t1p2_A5, the IC 100 is 50 mg/mL instead of 16 mg/mL; for antibody CnC2t1p1_B10, the IC 100 is >100 mg/mL instead of 12.5 mg/mL. For the latter antibody, binding characteristics were also corrected. As a consequence, the total number of neutralizing antibodies reported is 27 instead of 28. Changes affect Figures 3, 4, S3, S4, and S5, Tables S3 and S4, and text on pages 1, 3, 5, and 7. Importantly, the corrections have no impact on the conclusions in this paper. We apologize for any inconvenience that may have been caused by this error.

38The SARS-CoV-2 pandemic has unprecedented implications for public health, social 39 life, and world economy. Since approved drugs and vaccines are not available, new 40 options for COVID-19 treatment and prevention are highly demanded. To identify 41 SARS-CoV-2 neutralizing antibodies, we analysed the antibody response of 12 42 COVID-19 patients from 8 to 69 days post diagnosis. By screening 4,313 SARS-43CoV-2-reactive B cells, we isolated 255 antibodies from different time points as early 44 as 8 days post diagnosis. Among these, 28 potently neutralized authentic SARS-45CoV-2 (IC 100 as low as 0.04 µg/ml), showing a broad spectrum of V genes and low 46 levels of somatic mutations. Interestingly, potential precursors were identified in 47 naïve B cell repertoires from 48 healthy individuals that were sampled before the 48 COVID-19 pandemic. Our results demonstrate that SARS-CoV-2 neutralizing 49 antibodies are readily generated from a diverse pool of precursors, fostering the hope 50 of rapid induction of a protective immune response upon vaccination. 51 52