Development and evaluation of a serological test for diagnosis of COVID-19 with selected recombinant spike proteins

Abstract

Serological test is a valuable diagnostic tool for coronavirus disease 2019 (COVID-19). However, considerable improvements to these tests are needed, especially in the detection sensitivity. In this study, six recombinant nucleocapsid and spike proteins of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were prepared and evaluated, including three prokaryotic expression nucleocapsid proteins (rN, rN1, rN2) and three eukaryotic expression spike proteins (rS1, rS-RBD, rS-RBD-mFc). The recombinant proteins with the highest ELISA titers (rS1 and rS-RBD-mFc) were selected to develop a double-antigen sandwich colloidal gold immunochromatography assay (GICA) to detect total antibodies against SARS-CoV-2. The clinical evaluation results showed that the sensitivity and specificity of GICA were 92.09% (419/455) and 99.44% (706/710), respectively. Moreover, a significant number (65.63%, 21/32) of COVID-19 patients with undetectable viral RNA were correctly diagnosed by the GICA method. In conclusion, the eukaryotic expression spike proteins (rS1 and rS-RBD-mFc) are more suitable than the prokaryotic expression nucleocapsid proteins for serological diagnosis of SARS-CoV-2. The proposed GICA for detection of total antibodies could be a powerful complement to the current RNA tests for COVID-19.

Keywords: Colloidal gold immunochromatography assay (GICA); Recombinant protein; SARS-CoV-2; Serological diagnosis.

Fig. 1

Evaluation of the recombinant S and N proteins by indirect IgM-ELISA (a, c) and IgG-ELISA (b, d), respectively, with serum samples from seven healthy people and two COVID-19 patients (P1 and P2). Dashed lines refer to the cutoff values

Fig. 2

Schematic illustration (a), lower detection limit (b), and selectivity (c) of the sandwich-format GICA strip for total antibody detection against SARS-CoV-2