Detection of Nucleocapsid Antibody to SARS-CoV-2 is More Sensitive than Antibody to Spike Protein in COVID-19 Patients

Abstract

word count: 198 37 38for use under a CC0 license. ABSTRACT 39Background: SARS-CoV-2, the cause of coronavirus disease 2019 , is associated 40 with respiratory-related morbidity and mortality. Assays to detect virus-specific antibodies are 41 important to understand the prevalence of infection and the course of the immune response. 42 Methodology: Quantitative measurements of plasma or serum antibodies by luciferase 43 immunoprecipitation assay systems (LIPS) to the nucleocapsid and spike proteins were analyzed 44 in 100 cross-sectional or longitudinal samples from SARS-CoV-2-infected patients. A subset of 45 samples was tested with and without heat inactivation.46Results: Fifteen or more days after symptom onset, antibodies against SARS-CoV-2 47 nucleocapsid protein showed 100% sensitivity and 100% specificity, while antibodies to spike 48 protein were detected with 91% sensitivity and 100% specificity. Neither antibody levels nor the 49 rate of seropositivity were significantly reduced by heat inactivation of samples. Analysis of 50 daily samples from six patients with COVID-19 showed anti-nucleocapsid and spike antibodies 51 appearing between day 8 to day 14 after initial symptoms. Immunocompromised patients 52 generally had a delayed antibody response to SARS-CoV-2 compared to immunocompetent 53 patients. 54Conclusions: Antibody to the nucleocapsid protein of SARS-CoV-2 is more sensitive than 55 spike protein antibody for detecting early infection. Analyzing heat-inactivated samples by LIPS 56 is a safe and sensitive method for detecting SARS-CoV-2 antibodies. 57 58 feature of COVID-19 SARS-CoV-2 is virus-associated pneumonitis [5-7]. Unlike other highly 64 pathogenic coronaviruses such as SARS/SARS-CoV-1 and Middle East respiratory syndrome 65 coronavirus (MERS-CoV) [8], SARS-CoV-2 spreads more rapidly and reached six of the seven 66 continents, including North America [9], within three months of the initial outbreak. Nucleic 67 acid-based testing of oropharyngeal or nasopharyngeal swabs and saliva is useful for diagnosing 68 acute infection. SARS-CoV-2 virus RNA can often be detected in upper respiratory secretions at 69 the time of the first appearance of symptoms, peaks during the first week, and later declines with 70 time [10, 11]. RNA from SARS-CoV-2, like the related SARS-CoV-1 [12], can also be detected 71 in blood [11, 13], and high levels of circulating viral RNA are associated with more severe 72 disease [13]. 73 Assessment of the antibody response to SARS-CoV-2 should complement the RNA-74 based tests and improve our understanding of the disease course, contribute to epidemiological 75 studies and inform vaccine development. Antibodies to the nucleocapsid protein are the most 76 sensitive target for serologic diagnosis of infection with SARS-CoV-1 [14, 15]. Antibodies 77 against the spike protein of SARS-CoV-1, the target of neutralizing antibody and vaccine 78 development, emerge at a later time than those against the nucleocapsid protein. Recently, 79 several groups have reported serological dia...

Seroprevalence of antibodies against SARS-CoV-2 among health care workers in a large Spanish reference hospital

“…31,32 By increasing the number of viral antigens in our assay we may allow to increase its sensitivity, as responses to different antigens may present different kinetics and vary between individuals. 33 Nonetheless, we included determinations to three isotypes to capture a variety of responses between individuals and their relation to time from onset of symptoms. Also, their maintenance and role in protection are probably different.…”

A Systematic and Meta-Analysis Review on the Diagnostic Accuracy of Antibodies in the Serological Diagnosis of COVID-19.

“…One study did not specify the serological assay used and it was excluded from the review 28 . One study used a LIPS which is performed in solution, thus maintaining the native antigen conformation 29 .…”

“…We identi ed one study (Burbelo, 2020) reporting total antibody (Ab) based luciferase immunoprecipitation assay system (LIPS) using N and S antigens with sensitivities and speci cities of 0.91 (95 % CI 0.77-0.99) and 1.00 (0.80-1.00) and 1.00 (0.92-1.00) and 1.00 (0.92-1.00) respectively. We also identi ed studies reporting other Ab based serological assays and IgA based serological assays but results are not reported in this review.…”

Architecture and self-assembly of the SARS-CoV-2 nucleocapsid protein

“…One critical step in coronavirus replication is the assembly of the viral genomic RNA and nucleocapsid (N) protein into a ribonucleoprotein (RNP) complex, which in betacoronaviruses like SARS-CoV-2 is thought to form a helical filament structure that is packaged into virions through interactions with the membrane-spanning membrane (M) protein [23][24][25][26][27][28] . Despite its location within the viral particle rather than on its surface, patients infected with SARS-CoV-2 show higher and earlier antibody responses to the nucleocapsid protein than the surface spike protein 29,30 . As such, a better understanding of the SARS-CoV-2 N protein's structure, and structural differences between it and N proteins of related coronaviruses including SARS-CoV, may aid the development of sensitive and specific immunological tests.…”