tetano
Editor, Senior Moderator
Biosaf Health
. 2019 Dec;1(3):134-143.
doi: 10.1016/j.bsheal.2019.12.006. Epub 2019 Dec 20.
A Novel Luciferase Immunosorbent Assay Performs Better Than a Commercial Enzyme-Linked Immunosorbent Assay to Detect MERS-CoV Specific IgG in Humans and Animals
Wenling Wang[SUP] 1 [/SUP], Tianyu Wang[SUP] 1 2 [/SUP], Yao Deng[SUP] 1 [/SUP], Peihua Niu[SUP] 1 [/SUP], Ruhan A[SUP] 1 [/SUP], Jincun Zhao[SUP] 3 [/SUP], Malik Peiris[SUP] 4 [/SUP], Shixing Tang[SUP] 2 [/SUP], Wenjie Tan[SUP] 1 5 [/SUP]
Affiliations
Abstract
The Middle East respiratory syndrome (MERS) is a lethal zoonosis caused by MERS coronavirus (MERS-CoV) and poses a significant threat to public health worldwide. Therefore, a rapid, sensitive, and specific serologic test for detecting anti-MERS-CoV antibodies in both humans and animals is urgently needed for the successful management of this illness. Here, we evaluated various novel luciferase immunosorbent assays (LISA) based on nucleocapsid protein (NP) as well as fragments derived from spike protein (S) including subunit 1 (S1), N terminal domain (NTD), receptor-binding domain (RBD) and subunit 2 (S2) of S for the detection of MERS-CoV-specific IgG. Fusion proteins, including nanoluciferase (NLuc) and various fragments derived from the NP or S protein of MERS-CoV, were expressed in human embryonic kidney 293 T cells. LISAs that detected anti-MERS-CoV IgG were further developed using cell lysates expressing various fusion proteins. Panels of human or animal samples infected with MERS-CoV were used to analyze the sensitivity and specificity of various LISAs in reference to a MERS-CoV RT-PCR, commercial S1-based ELISA, and pseudovirus particle neutralization test (ppNT). Our results showed that the S1-, RBD-, and NP-LISAs were more sensitive than the NTD- and S2-LISAs for the detection of anti-MERS-CoV IgG. Furthermore, the S1-, RBD-, and NP-LISAs were more sensitive (by at least 16-fold) than the commercially available S1-ELISA. Moreover, the S1-, RBD-, and NP-LISA specifically recognized anti-MERS-CoV IgG and did not cross-react with samples derived from other human CoV (OC43, 229E, HKU1, NL63)-infected patients. More importantly, these LISAs proved their applicability and reliability for detecting anti-MERS-CoV IgG in samples from camels, monkeys, and mice, among which the RBD-LISA exhibited excellent performance. The results of this study suggest that the novel MERS-CoV RBD- and S1- LISAs are highly effective platforms for the rapid and sensitive detection of anti-MERS-CoV IgG in human and animal samples. These assays have the potential to be used as serologic tests for the management and control of MERS-CoV infection.
Keywords: Luciferase immunosorbent assay (LISA); MERS-CoV; Samples of humans and animals; Serological IgG detection.
. 2019 Dec;1(3):134-143.
doi: 10.1016/j.bsheal.2019.12.006. Epub 2019 Dec 20.
A Novel Luciferase Immunosorbent Assay Performs Better Than a Commercial Enzyme-Linked Immunosorbent Assay to Detect MERS-CoV Specific IgG in Humans and Animals
Wenling Wang[SUP] 1 [/SUP], Tianyu Wang[SUP] 1 2 [/SUP], Yao Deng[SUP] 1 [/SUP], Peihua Niu[SUP] 1 [/SUP], Ruhan A[SUP] 1 [/SUP], Jincun Zhao[SUP] 3 [/SUP], Malik Peiris[SUP] 4 [/SUP], Shixing Tang[SUP] 2 [/SUP], Wenjie Tan[SUP] 1 5 [/SUP]
Affiliations
- PMID: 32501446
- PMCID: PMC7148641
- DOI: 10.1016/j.bsheal.2019.12.006
Abstract
The Middle East respiratory syndrome (MERS) is a lethal zoonosis caused by MERS coronavirus (MERS-CoV) and poses a significant threat to public health worldwide. Therefore, a rapid, sensitive, and specific serologic test for detecting anti-MERS-CoV antibodies in both humans and animals is urgently needed for the successful management of this illness. Here, we evaluated various novel luciferase immunosorbent assays (LISA) based on nucleocapsid protein (NP) as well as fragments derived from spike protein (S) including subunit 1 (S1), N terminal domain (NTD), receptor-binding domain (RBD) and subunit 2 (S2) of S for the detection of MERS-CoV-specific IgG. Fusion proteins, including nanoluciferase (NLuc) and various fragments derived from the NP or S protein of MERS-CoV, were expressed in human embryonic kidney 293 T cells. LISAs that detected anti-MERS-CoV IgG were further developed using cell lysates expressing various fusion proteins. Panels of human or animal samples infected with MERS-CoV were used to analyze the sensitivity and specificity of various LISAs in reference to a MERS-CoV RT-PCR, commercial S1-based ELISA, and pseudovirus particle neutralization test (ppNT). Our results showed that the S1-, RBD-, and NP-LISAs were more sensitive than the NTD- and S2-LISAs for the detection of anti-MERS-CoV IgG. Furthermore, the S1-, RBD-, and NP-LISAs were more sensitive (by at least 16-fold) than the commercially available S1-ELISA. Moreover, the S1-, RBD-, and NP-LISA specifically recognized anti-MERS-CoV IgG and did not cross-react with samples derived from other human CoV (OC43, 229E, HKU1, NL63)-infected patients. More importantly, these LISAs proved their applicability and reliability for detecting anti-MERS-CoV IgG in samples from camels, monkeys, and mice, among which the RBD-LISA exhibited excellent performance. The results of this study suggest that the novel MERS-CoV RBD- and S1- LISAs are highly effective platforms for the rapid and sensitive detection of anti-MERS-CoV IgG in human and animal samples. These assays have the potential to be used as serologic tests for the management and control of MERS-CoV infection.
Keywords: Luciferase immunosorbent assay (LISA); MERS-CoV; Samples of humans and animals; Serological IgG detection.