SARS-CoV-2 N-antigenemia: A new COVID-19 marker and a potential alternative to nucleic acid amplification techniques

Background Molecular assays on nasopharyngeal swabs remain the cornerstone of COVID-19 diagnostic. Despite massive worldwide efforts, the high technicalities of nasopharyngeal sampling and molecular assays, as well as scarce resources of reagents, limit our testing capabilities. Several strategies failed, to date, to fully alleviate this testing process (e.g. saliva sampling or antigen testing on nasopharyngeal samples). We assessed the performances of a new ELISA microplate assay quantifying SARS-CoV-2 nucleocapsid antigen (N-antigen) in serum or plasma. Methods The specificity of the assay, determined on 63 non-COVID patients, was 98.4% (95% confidence interval [CI], 85.3 to 100). Performances were determined on 227 serum samples from 165 patients with RT-PCR confirmed SARS-CoV-2 infection included in the French COVID and CoV-CONTACT cohorts. Findings Sensitivity was 132/142, 93.0% (95% CI, 84.7 to 100), within the first two weeks after symptoms onset. A subset of 73 COVID-19 patients had a serum collected within 24 hours following or preceding a positive nasopharyngeal swab. Among patients with high nasopharyngeal viral loads, Ct value below 30 and 33, only 1/50 and 4/67 tested negative for N-antigenemia, respectively. Among patients with a negative nasopharyngeal RT-PCR, 8/12 presented positive N-antigenemia. The lower respiratory tract was explored for 6/8 patients, showing positive PCR in 5 cases. Interpretation This is the first demonstration of the N-antigen antigenemia during COVID-19. Its detection presented a robust sensitivity, especially within the first 14 days after symptoms onset and high nasopharyngeal viral loads. These findings have to be confirmed with higher representation of outpatients. This approach could provide a valuable new option for COVID-19 diagnosis, only requiring a blood draw and easily scalable in all clinical laboratories. ### Competing Interest Statement Dr. Visseaux reports grants, personal fees and non-financial support from Qiagen, personal fees and non-financial support from BioMerieux, personal fees from Hologic, outside the submitted work. ### Clinical Trial NCT04262921, [NCT04259892][1] ### Funding Statement This study has been funded in part by the REACTing (REsearch & ACTion emergING infectious diseases) consortium, by a grant of the French Ministry of Health (PHRC #20-0424) and the ANRS (Agence Nationale de la Recherche sur le SIDA et les hepatites virales). The study was supported by AAZ (Boulogne-Billancourt, France) in the form of free consumables and they had no role in the study conception, design, conduct, data analysis or manuscript preparation. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: COVID patients are included in the French COVID (clinicaltrials.gov [NCT04262921][2]) and CoV-CONTACT cohorts (clinicaltrials.gov [NCT04259892][1]). Ethics approval was given by the French Ethics Committee CPP-Ile-de-France 6 (ID RCB: 2020-A00256-33 and ID RCB: 2020-A00280-39) and the French National Data Protection Commission (approval #920102). All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes A file compiling all data used in this article is available on Mendeley Data public repository (). [1]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT04259892&atom=%2Fmedrxiv%2Fearly%2F2020%2F09%2F15%2F2020.09.14.20191759.atom [2]: /lookup/external-ref?link_type=CLINTRIALGOV&access_num=NCT04262921&atom=%2Fmedrxiv%2Fearly%2F2020%2F09%2F15%2F2020.09.14.20191759.atom