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Open Access 01-12-2022 | SARS-CoV-2 | Methodology

Preparation of the luciferase-labeled antibody for improving the detection sensitivity of viral antigen

Authors: Ying Tang, Yuchang Li, Sen Zhang, Jing Li, Yi Hu, Wenguang Yang, Yuehong Chen, Chengfeng Qin, Tao Jiang, Xiaoping Kang

Published in: Virology Journal | Issue 1/2022

Abstract

Background

Viral antigen detection test is the most common method used to detect viruses in the field rapidly. However, due to the low sensitivity, it can only be used as an auxiliary diagnosis method for virus infection. Improving sensitivity is crucial for developing more accurate viral antigen tests. Nano luciferase (Nluc) is a sensitive reporter that has not been used in virus detection.

Results

In this study, we produced an intracellularly Nluc labeled detection antibody (Nluc-ch2C5) and evaluated its ability to improve the detection sensitivity of respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens. Compared with the traditional horse-radish peroxidase (HRP) labeled antibody (HRP-ch2C5), Nluc-ch2C5 was 41 times more sensitive for inactivated SARS-CoV-2 virus by sandwich chemiluminescence ELISA. Then we applied Nluc-ch2C5 to establish an automatic magnet chemiluminescence immune assay (AMCA) for the SARS-CoV-2 viral spike protein, the limit of detection was 68 pfu/reaction. The clinical sensitivity and specificity reached 75% (24/32) and 100% (48/48) using 32 PCR-positive and 48 PCR-negative swab samples for clinical evaluation, which is more sensitive than the commercial ELSA kit and colloid gold strip kit.

Conclusions

Here, monoclonal antibody ch2C5 served as a model antibody and the SARS-CoV-2 served as a model pathogen. The Nluc labeled detecting antibody (Nluc-ch2C5) significantly improved the detection sensitivity of SARS-CoV-2 antigen. This labeling principle applies to other viral infections, so this labeling and test format could be expected to play an important role in detecting other virus antigens.

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Edridge AWD, van der Hoek L. Emerging orthobunya viruses associated with CNS disease. PLoS Negl Trop Dis. 2020;14(10): e0008856. https://doi.org/10.1371/journal.pntd.0008856.CrossRefPubMedPubMedCentral

Malvy D, McElroy AK, de Clerck H, Günther S, van Griensven J. Ebola virus disease. Lancet. 2019;393:936–48. https://doi.org/10.1016/S0140-6736(18)33132-5.CrossRefPubMed

Wang C, Horby PW, Hayden FG, Gao GF. A novel coronavirus outbreak of global health concern. Lancet. 2020;395:470–3.CrossRef

Wang H, Wang W, Tan W. Summary of the detection kits for SARS-CoV-2 approved by the national medical products administration of china and their application for diagnosis of COVID-19. Virol Sin. 2020;35(6):699–712. https://doi.org/10.1007/s12250-020-00331-1.CrossRefPubMedPubMedCentral

Onozuka N, Ohki T, Oka N, Maoka T. One-step real-time multiplex reverse transcription-polymerase chain reaction assay with melt curve analysis for detection of potato leafroll virus, potato virus S, potato virus X, and potato virus Y. Virol J. 2021;18(1):131. https://doi.org/10.1186/s12985-021-01591-3.PMID:34187522;PMCID:PMC8243585.CrossRefPubMedPubMedCentral

Gdoura M, Abouda I, Mrad M, et al. SARS-CoV2 RT-PCR assays: In vitro comparison of 4 WHO approved protocols on clinical specimens and its implications for real laboratory practice through variant emergence. Virol J. 2022;19:54.CrossRef

Li Y, Li J, Zhang Y, Dai L, Li L, Liu J, Zhang S, Wu X, Hu Y, Qin C, Jiang T, Kang X. Development of an automatic integrated gene detection system for novel severe acute respiratory syndrome-related coronavirus (SARS-CoV2). Emerg Microbes Infect. 2020;9(1):1489–96. https://doi.org/10.1080/22221751.2020.1782774.PMID:32543298;PMCID:PMC7473122.CrossRefPubMedPubMedCentral

Tang Y, Wang Y, Li Y, Zhao H, Zhang S, Zhang Y, Li J, Chen Y, Wu X, Qin C, Jiang T, Kang X. An integrated rapid nucleic acid detection assay based on recombinant polymerase amplification for SARS-CoV-2. Virol Sin. 2022;37(1):138–41. https://doi.org/10.1016/j.virs.2022.01.006.CrossRefPubMedPubMedCentral

Fabiani L, Saroglia M, Galatà G, De Santis R, Fillo S, Luca V, Faggioni G, D’Amore N, Regalbuto E, Salvatori P, Terova G, Moscone D, Lista F, Arduini F. Magnetic beads combined with carbon black-based screen-printed electrodes for COVID-19: a reliable and miniaturized electrochemical immunosensor for SARS-CoV-2 detection in saliva. Biosens Bioelectron. 2021;1(171): 112686. https://doi.org/10.1016/j.bios.2020.112686.CrossRef

10.

Zhang X, Zhu H, Zheng X, et al. A double-antibody sandwich ELISA for sensitive and specific detection of swine fibrinogen-like protein 1[J]. Front Immunol. 2021;12:1290.

11.

Grandien M. Viral diagnosis by antigen detection techniques[J]. Clin Diagn Virol. 1996;5(2–3):81–90.CrossRef

12.

Mak GCK, Lau SSY, Wong KKY, Chow NLS, Lau CS, Lam ETK, Chan RCW, Tsang DNC. Evaluation of rapid antigen detection kit from the WHO Emergency Use List for detecting SARS-CoV-2. J Clin Virol. 2021;134:104712. https://doi.org/10.1016/j.jcv.2020.104712.CrossRefPubMed

13.

Hall MP, Unch J, Binkowski BF, Valley MP, Butler BL, Wood MG, Otto P, Zimmerman K, Vidugiris G, Machleidt T, Robers MB, Benink HA, Eggers CT, Slater MR, Meisenheimer PL, Klaubert DH, Fan F, Encell LP, Wood KV. Engineered luciferase reporter from a deep sea shrimp utilizing a novel imidazopyrazinone substrate. ACS Chem Biol. 2012;7(11):1848–57. https://doi.org/10.1021/cb3002478.CrossRefPubMedPubMedCentral

14.

Widder EA. Bioluminescence in the ocean: origins of biological, chemical, and ecological diversity. Science. 2010;328(5979):704–8. https://doi.org/10.1126/science.1174269.CrossRefPubMed

15.

Li YC, Hu Y, Wu XY, Huo NF, Li J, Zhang S, Jiang T, Kang XP. A sensitive nano luciferase immune complex assay system for highly sensitive and specific detection of antibodies against tick-borne encephalitis virus. Vector Borne Zoonotic Dis. 2019;19(5):365–9. https://doi.org/10.1089/vbz.2018.2330.CrossRefPubMed

16.

Shi Y, Wang G, Cai XP, Deng JW, Zheng L, Zhu HH, Zheng M, Yang B, Chen Z. An overview of COVID-19. J Zhejiang Univ Sci B. 2020;21(5):343–60. https://doi.org/10.1631/jzus.B2000083.CrossRefPubMedPubMedCentral

17.

Rusanen J, Kareinen L, Szirovicza L, Ugurlu H, Levanov L, Jääskeläinen A, Ahava M, Kurkela S, Saksela K, Hedman K, Vapalahti O, Hepojoki J. A generic, scalable, and rapid time-resolved Förster resonance energy transfer-based assay for antigen detection— SARS-CoV-2 as a proof of concept. MBio. 2021;12:e00902-e921.CrossRef

18.

Fabiani L, Saroglia M, Galatà G, De Santis R, Fillo S, Luca V, Faggioni G, D’Amore N, Regalbuto E, Salvatori P, Terova G, Moscone D, Lista F, Arduini F. Magnetic beads combined with carbon black-based screen-printed electrodes for COVID-19: a reliable and miniaturized electrochemical immunosensor for SARS-CoV-2 detection in saliva. Biosens Bioelectron. 2021;171: 112686. https://doi.org/10.1016/j.bios.2020.112686.CrossRefPubMed

19.

Jiang W, Wang J, Jiao S, Chenjian G, Wei X, Chen B, Wang R, Chen H, Xie Y, Wang A, Li G, Zeng D, Zhang J, Zhang M, Wang S, Wang M, Gui X. Characterization of MW06, a human monoclonal antibody with cross-neutralization activity against both SARS-CoV-2 and SARS-CoV. MAbs. 2021. https://doi.org/10.1080/19420862.2021.1953683.CrossRefPubMedPubMedCentral

20.

Li Y, Wang M, Wu H, et al. Development of a rapid neutralizing antibody test for SARS-CoV-2 and its application for neutralizing antibody screening and vaccinated serum testing. Infect Med. 2022. https://doi.org/10.1016/j.imj.2022.04.003.CrossRef

21.

Li JF, He L, Deng YQ, Qi SH, Chen YH, Zhang XL, Hu SX, Fan RW, Zhao GY, Qin CF. Generation and characterization of a nanobody against SARS-CoV. Virol Sin. 2021;36(6):1484–91. https://doi.org/10.1007/s12250-021-00436-1.CrossRefPubMedPubMedCentral

22.

Berg EA, Fishman JB. Labeling antibodies [J]. Cold Spring Harbor Protocols, 2020(7):pdb. top099242.

23.

Berg EA, Fishman JB. Labeling antibodies using NHS-fluorescein. Cold Spring Harb Protoc. 2019. https://doi.org/10.1101/pdb.prot099283.CrossRefPubMed

24.

Berg EA, Fishman JB. Labeling antibodies using a maleimido dye. Cold Spring Harb Protoc. 2019. https://doi.org/10.1101/pdb.prot099291.CrossRefPubMed

25.

Chang L, Rissin DM, Fournier DR, Piech T, Patel PP, Wilson DH, Duffy DC. Single molecule enzyme-linked immunosorbent assays: theoretical considerations. J Immunol Methods. 2012;30;378(1–2):102–15. https://doi.org/10.1016/j.jim.2012.02.011.CrossRef

26.

Majdinasab M, Zareian M, Zhang Q, Li P. Development of a new format of competitive immunochromatographic assay using secondary antibody-europium nanoparticle conjugates for ultrasensitive and quantitative determination of ochratoxin A. Food Chem. 2019;1(275):721–9. https://doi.org/10.1016/j.foodchem.2018.09.112 (Epub 2018 Sep 19 PMID: 30724255).CrossRef

27.

Wijesuriya SD, Pongo E, Tomic M, Zhang F, Garcia-Rodriquez C, Conrad F, Farr-Jones S, Marks JD, Horwitz AH. Antibody engineering to improve manufacturability. Protein Expr Purif. 2018;149:75–83. https://doi.org/10.1016/j.pep.2018.04.003 (Epub 2018 Apr 12 PMID: 29655788).CrossRefPubMed

28.

König J, Hust M, van den Heuvel J. Validation of the production of antibodies in different formats in the HEK 293 transient gene expression system. Methods Mol Biol. 2021;2247:59–76. https://doi.org/10.1007/978-1-0716-1126-5_4 (PMID: 33301112).CrossRefPubMed

29.

Adams JJ, Sidhu SS. Synthetic antibody technologies. Curr Opinion Struct Biol. 2014;21:1–9.CrossRef

30.

Seeliger D, et al. Boosting antibody developability through rational sequence optimization. MAbs. 2015;7(3):505–15.

Title: Preparation of the luciferase-labeled antibody for improving the detection sensitivity of viral antigen
Authors: Ying Tang
Yuchang Li
Sen Zhang
Jing Li
Yi Hu
Wenguang Yang
Yuehong Chen
Chengfeng Qin
Tao Jiang
Xiaoping Kang
Publication date: 01-12-2022
Publisher: BioMed Central
Keyword: SARS-CoV-2
Published in: Virology Journal / Issue 1/2022
Electronic ISSN: 1743-422X
DOI: https://doi.org/10.1186/s12985-022-01855-6

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