A Sensitive Method for Detecting Zika Virus Antigen in Patients’ Whole-Blood Specimens as an Alternative Diagnostic Approach

Lum, Fok‐Moon; Lin, Cui; Susova, Olga Yu.; Teo, Teck‐Hui; Fong, Siew‐Wai; Mak, Tze Minn; Lee, Linda Kay; Chong, Chia-Yin; Lye, David C.; Lin, Raymond Tzer‐Pin; Merits, Andres; Leo, Yee‐Sin; Ng, Lisa F. P.

doi:10.1093/infdis/jix276

Cited by 25 publications

(30 citation statements)

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“…Anti-ZIKV IgM and IgG levels of ZIKV patients from the Singapore outbreak in 2016 [28][29][30] were longitudinally assessed using virion-based ELISA. 18,[24][25][26][27] The majority of the patients showed a robust ZIKVspecific humoral response (Figure 1a IgG isotypes produced by ZIKV patients were then determined, and highest titres of anti-ZIKV IgG1 and IgG3 subtypes were produced at early convalescent for IgG3 and late convalescent for IgG1 (Figure 1d).…”

Section: Zikv Patients Produce a Robust And Protective Humoral Responsementioning

confidence: 99%

“…After 2 h, media were removed and Dulbecco's modified Eagle's medium (DMEM; GE Healthcare Life Sciences, Pittsburgh, PA, USA) with 10% foetal bovine serum (FBS; GE Healthcare Life Sciences) was added. After 48 h, cells were harvested and stained as described, 54 using ZIKV NS3 protein-specific rabbit polyclonal antibody 29 or DENV human monoclonal antibody 1B, 25 and counter-stained with fluorophoretagged goat anti-rabbit or anti-human IgG (H+L) (Thermo Fisher Scientific). Cells were acquired with MACSQuant Analyser 10 (Miltenyi Biotec, Bergisch Gladbach, Germany).…”

Section: Sero-neutralisationmentioning

confidence: 99%

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Novel differential linear B‐cell epitopes to identify Zika and dengue virus infections in patients

et al. 2019

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Objectives Recent Zika virus (ZIKV) outbreaks challenged existing laboratory diagnostic standards, especially for serology‐based methods. Because of the genetic and structural similarity of ZIKV with other flaviviruses, this results in cross‐reactive antibodies, which confounds serological interpretations. Methods Plasma from Singapore ZIKV patients was screened longitudinally for antibody responses and neutralising capacities against ZIKV. Samples from healthy controls, ZIKV patients and DENV patients were further assessed using ZIKV and DENV peptides of precursor membrane (prM), envelope (E) or non‐structural 1 (NS1) viral proteins in a peptide‐based ELISA for epitope identification. Identified epitopes were re‐validated and diagnostically evaluated using sera of patients with DENV, bacteria or unknown infections from Thailand. Results Long‐lasting ZIKV‐neutralising antibodies were elicited during ZIKV infection. Thirteen potential linear B‐cell epitopes were identified, and of these, four common flavivirus, three ZIKV‐specific and one DENV‐specific differential epitopes had more than 50% sensitivity and specificity. Notably, ZIKV‐specific peptide 26 on domain I/II of E protein (amino acid residues 271–288) presented 80% sensitivity and 85.7% specificity. Importantly, the differential epitopes also showed significance in differentiating non‐flavivirus patient samples. Conclusion Linear B‐cell epitope candidates to differentiate between ZIKV and DENV infections were identified, providing the first step towards the design of a much‐needed serology‐based assay.

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Section: Zikv Patients Produce a Robust And Protective Humoral Responsementioning

confidence: 99%

Section: Sero-neutralisationmentioning

confidence: 99%

Novel differential linear B‐cell epitopes to identify Zika and dengue virus infections in patients

et al. 2019

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“…Anti-ZIKV IgM and IgG levels of ZIKV patients from the Singapore outbreak in 2016 [24,25,38] were longitudinally assessed using virion-based ELISA [18,32,34–36]. Majority of the patients showed a robust ZIKV-specific humoral response (Figures 1A-C, Supplementary Figure 1C).…”

Section: Resultsmentioning

confidence: 99%

“…After 2 h, media were removed and Dulbecco’s Modified Eagle Medium (DMEM; HyClone) with 10% fetal bovine serum (FBS; HyClone) were added. After 48 h, cells were harvested and stained as described [37], using ZIKV NS3 protein-specific rabbit polyclonal antibody [38] or DENV human monoclonal antibody 1B [34], and counter-stained with fluorophore-tagged goat anti-rabbit or anti-human IgG (H+L) (Life Technologies) respectively. Cells were acquired with MacsQuant Analyzer 10 (Miltenyi-Biotec).…”

Section: Methodsmentioning

confidence: 99%

Novel differential linear B-cell epitopes to identify Zika and dengue virus infections in patients

Amrun

Yee

Bakar

et al. 2019

Preprint

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word count: 200 words 35 Text word count: 3612 words 36 Abstract (200 words) 37 Background 38 Recent Zika virus (ZIKV) outbreaks challenged existing laboratory diagnostic 39 standards, especially for serology-based methods. Due to the genetic and structural 40 similarity of ZIKV with other flaviviruses, this results in cross-reactive antibodies 41 which confounds serological interpretations. 42 43 Methods 44 Plasma from Singapore ZIKV patients was screened longitudinally for antibody 45 responses and neutralizing capacities against ZIKV. Samples from healthy controls, 46ZIKV and DENV patients were further assessed using ZIKV and DENV peptides of 47 precursor membrane (prM), envelope (E) or non-structural 1 (NS1) viral proteins in a 48 peptide-based ELISA for epitope identification. Identified epitopes were re-validated 49 and diagnostically evaluated using sera of patients with DENV, bacteria or unknown 50 infections from Thailand. 51 52 Results 53Long-lasting ZIKV-neutralizing antibodies were elicited during ZIKV infection. 54Thirteen potential linear B-cell epitopes were identified and of these, four common 55 flavivirus, three ZIKV-specific, and one DENV-specific differential epitopes had more 56 than 50% sensitivities and specificities. Notably, ZIKV-specific peptide 26 on domain 57 I/II of E protein (amino acid residues 271-288) presented 80% sensitivity and 85.7% 58 specificity. Importantly, the differential epitopes also showed significance in 59 differentiating non-flavivirus patient samples. 60 61 Conclusions 62 Linear B-cell epitope candidates to differentiate ZIKV and DENV infections were 63 identified, providing the first step towards the design of a much-needed serology-64 based assay. 65 66 Methods 97 Ethics statement 98 Written informed consent was obtained from participants in accordance with the 99 tenets of the Declaration of Helsinki. Study protocols of Singapore ZIKV (2016-2018) 100 and DENV (2010-2012) patient cohorts were approved by the SingHealth 101 Centralized Institutional Review Board (CIRB Ref: 2016/2219) and National 102 Healthcare Group (NHG) Domain Specific Review Board (DSRB-E-2009/432) 103 respectively. Specimens from Singapore healthy donors (2010-2015) and patients 104 from Thailand (2011-2013) were collected in accordance to study guidelines of 105 approval numbers: NUS-IRB 09-256 and NUS-IRB 10-445; MUTM 2011-008-01, 106 OXTREC 42-10 and TCAB-01-11 respectively. 107 108 Study subjects and sample collection 109 Singapore ZIKV patients 110Collection of specimens from subjects during the ZIKV outbreak in 2016 was 111 previously described [24]. Briefly, 65 patients that were RT-PCR positive for ZIKV in 112 whole blood or urine, and negative for DENV RT-PCR were enrolled [25]. Whole 113 blood specimens were collected in EDTA-coated vacutainer tubes (Becton 114 Dickinson) after peripheral venipuncture and were centrifuged at 12000 rpm for 10 115 min. Plasma was collected and heat-inactivated for 30 min at 56°C before storage at 116 -80°C. Specimens were obtained over a period of si...

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“…After washing with TBS-T, the tissues were then treated with 10% goat serum for 30 min to block the nonspecific reaction. The tissues were then incubated overnight with various combinations of a ZIKV in-house antibody [13] (1:100) and CD 163 (Thermofisher; 1:50) at 4°C. The slides were incubated with secondary antibodies (1:500) such as Alexa Fluor ® 488 goat anti rabbit IgG (Life technologies) and Alexa Fluor ® 594 goat anti mouse IgG (Invitrogen) for 30 min in the dark, and the nuclei were counter stained with Vectashield™ Hard Set mounting medium with DAPI.…”

Section: Methodsmentioning

confidence: 99%

Trimester-specific Zika virus infection affects placental responses in women

Lum

Narang

Hue

et al. 2019

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A Sensitive Method for Detecting Zika Virus Antigen in Patients’ Whole-Blood Specimens as an Alternative Diagnostic Approach

Abstract: SummaryThe reemergence of Zika virus (ZIKV) warrants the need to develop complementary diagnostic measures. In this study, flow cytometry was used to determine the presence of ZIKV NS3 antigen in blood monocytes from ZIKV-infected patients

Cited by 25 publications

References 30 publications

Novel differential linear B‐cell epitopes to identify Zika and dengue virus infections in patients

Novel differential linear B‐cell epitopes to identify Zika and dengue virus infections in patients

Novel differential linear B-cell epitopes to identify Zika and dengue virus infections in patients

Trimester-specific Zika virus infection affects placental responses in women

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