TY - JOUR
T1 - Simultaneous detection of antibody responses to multiple SARS-CoV-2 antigens by a Western blot serological assay
AU - Hsiao, Chia Chun
AU - Chiang, Yi Wei
AU - Chao, Tai Ling
AU - Tsai, Zen Uong
AU - Wang, Ting Xuan
AU - Jiang, Yu Wei
AU - Hsu, Hsiang Fu
AU - Lu, De Chao
AU - Wang, Jann Tay
AU - Wang, Jen Ren
AU - Wang, An Bang
AU - Chang, Sui Yuan
AU - Chang, Shih Chung
N1 - Funding Information:
This work was supported by the Ministry of Science and Technology, Taiwan (grant numbers MOST111-2320-B-002–047, MOST111-2221-E-002–199, MOST110-2311-B-002–010, MOST110-2221-E-002–085, and MOST109-2327-B-002–009) and National Taiwan University (grant numbers 109L883703, 110L880802, and 111L880802).
Publisher Copyright:
© 2022, The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.
PY - 2022/12
Y1 - 2022/12
N2 - Abstract: The nucleic acid test is still the standard assessment for the diagnosis of coronavirus disease 2019 (COVID-19), which is caused by human infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In addition to supporting the confirmation of disease cases, serological assays are used for the analysis of antibody status and epidemiological surveys. In this study, a single Western blot strip (WBS) coated with multiple Escherichia coli (E. coli)-expressed SARS-CoV-2 antigens was developed for comprehensive studies of antibody profiles in COVID-19 patient sera. The levels of specific antibodies directed to SARS-CoV-2 spike (S), S2, and nucleocapsid (N) proteins were gradually increased with the same tendency as the disease progressed after hospitalization. The signal readouts of S, S2, and N revealed by the multi-antigen-coated WBS (mWBS)-based serological assay (mWBS assay) also demonstrated a positive correlation with the SARS-CoV-2 neutralizing potency of the sera measured by the plaque reduction neutralization test (PRNT) assays. Surprisingly, the detection signals against the unstructured receptor-binding domain (RBD) purified from E. coli inclusion bodies were not observed, although the COVID-19 patient sera exhibited strong neutralizing potency in the PRNT assays, suggesting that the RBD-specific antibodies in patient sera mostly recognize the conformational epitopes. Furthermore, the mWBS assay identified a unique and major antigenic epitope at the residues 1148, 1149, 1152, 1155, and 1156 located within the 1127–1167 fragment of the S2 subunit, which was specifically recognized by the COVID-19 patient serum. The mWBS assay can be finished within 14–16 min by using the automatic platform of Western blotting by thin-film direct coating with suction (TDCS WB). Collectively, the mWBS assay can be applied for the analysis of antibody responses, prediction of the protective antibody status, and identification of the specific epitope. Key points: • A Western blot strip (WBS) coated with multiple SARS-CoV-2 antigens was developed for the serological assay. • The multi-antigen-coated WBS (mWBS) can be utilized for the simultaneous detection of antibody responses to multiple SARS-CoV-2 antigens. • The mWBS-based serological assay (mWBS assay) identified a unique epitope recognized by the COVID-19 patient serum.
AB - Abstract: The nucleic acid test is still the standard assessment for the diagnosis of coronavirus disease 2019 (COVID-19), which is caused by human infection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In addition to supporting the confirmation of disease cases, serological assays are used for the analysis of antibody status and epidemiological surveys. In this study, a single Western blot strip (WBS) coated with multiple Escherichia coli (E. coli)-expressed SARS-CoV-2 antigens was developed for comprehensive studies of antibody profiles in COVID-19 patient sera. The levels of specific antibodies directed to SARS-CoV-2 spike (S), S2, and nucleocapsid (N) proteins were gradually increased with the same tendency as the disease progressed after hospitalization. The signal readouts of S, S2, and N revealed by the multi-antigen-coated WBS (mWBS)-based serological assay (mWBS assay) also demonstrated a positive correlation with the SARS-CoV-2 neutralizing potency of the sera measured by the plaque reduction neutralization test (PRNT) assays. Surprisingly, the detection signals against the unstructured receptor-binding domain (RBD) purified from E. coli inclusion bodies were not observed, although the COVID-19 patient sera exhibited strong neutralizing potency in the PRNT assays, suggesting that the RBD-specific antibodies in patient sera mostly recognize the conformational epitopes. Furthermore, the mWBS assay identified a unique and major antigenic epitope at the residues 1148, 1149, 1152, 1155, and 1156 located within the 1127–1167 fragment of the S2 subunit, which was specifically recognized by the COVID-19 patient serum. The mWBS assay can be finished within 14–16 min by using the automatic platform of Western blotting by thin-film direct coating with suction (TDCS WB). Collectively, the mWBS assay can be applied for the analysis of antibody responses, prediction of the protective antibody status, and identification of the specific epitope. Key points: • A Western blot strip (WBS) coated with multiple SARS-CoV-2 antigens was developed for the serological assay. • The multi-antigen-coated WBS (mWBS) can be utilized for the simultaneous detection of antibody responses to multiple SARS-CoV-2 antigens. • The mWBS-based serological assay (mWBS assay) identified a unique epitope recognized by the COVID-19 patient serum.
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U2 - 10.1007/s00253-022-12288-0
DO - 10.1007/s00253-022-12288-0
M3 - Article
C2 - 36404356
AN - SCOPUS:85142251149
SN - 0175-7598
VL - 106
SP - 8183
EP - 8194
JO - Applied Microbiology and Biotechnology
JF - Applied Microbiology and Biotechnology
IS - 24
ER -