Development and evaluation of a low cost IgG ELISA test based in RBD protein for COVID-19

Villafañe, Luciana; Vaulet, Lucía Gallo; Viere, Florencia M.; Klepp, Laura I.; Forrellad, Marina Andrea; Bigi, María Mercedes; Romanó, Maria; Magistrelli, Giovanni; Fermepin, Marcelo Rodríguez; Bigi, Fabiana

doi:10.1016/j.jim.2021.113182

Cited by 12 publications

(8 citation statements)

References 14 publications

(24 reference statements)

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“…During the validation process, the optimized iELISA exhibited high sensitivity and specificity values, in line with some previously reported works (or even better) (Ibrahim et al 2021 ; Mehdi et al 2021 ; Ojeda et al 2021 ; Villafañe et al 2022 ; Ayón-Núñez et al 2022 ; Amanat et al 2020 ). However, to make a correct comparison between the performance of different methods, it is necessary to have additional information (such as timing, symptoms), which sometimes is not provided by all the studies.…”

Section: Discussionsupporting

confidence: 88%

Design and optimization of an IgG human ELISA assay reactive to recombinant RBD SARS-CoV-2 protein

Rodríguez

Ceaglio

Gugliotta

et al. 2022

Appl Microbiol Biotechnol

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Serology assays are essential tools to mitigate the effect of COVID-19, help to identify previous SARS-CoV-2 infections or vaccination, and provide data for surveillance and epidemiologic studies. In this study, we report the production and purification process of the receptor-binding domain (RBD) of SARS-CoV-2 in HEK293 cells, which allowed the design, optimization, and validation of an indirect ELISA (iELISA) for the detection of human anti-RBD antibodies. To find the optimal conditions of this iELISA, a multivariate strategy was performed throughout design of experiments (DoE) and response surface methodology (RSM), one of the main tools of quality by design (QbD) approach. The adoption of this strategy helped to reduce the time and cost during the method development stage and to define an optimum condition within the analyzed design region. The assay was then validated, exhibiting a sensitivity of 94.24 (86.01–98.42%; 95% CI) and a specificity of 95.96% (89.98–98.89%; 95% CI). Besides, the degree of agreement between quality results assessed using kappa’s value was 0.92. Hence, this iELISA represents a high-throughput technique, simple to perform, reliable, and feasible to be scaled up to satisfy the current demands. Since RBD is proposed as the coating antigen, the intended use of this iELISA is not only the detection of previous exposure to the virus, but also the possibility of detecting protective immunity. Key points • RBD was produced in 1-L bioreactor and highly purified. • An iELISA assay was optimized applying QbD concepts. • The validation procedure demonstrated that this iELISA is accurate and precise. Graphical Abstract Supplementary Information The online version contains supplementary material available at 10.1007/s00253-022-12254-w.

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Section: Discussionsupporting

confidence: 88%

Design and optimization of an IgG human ELISA assay reactive to recombinant RBD SARS-CoV-2 protein

Rodríguez

Ceaglio

Gugliotta

et al. 2022

Appl Microbiol Biotechnol

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“…Our optimized ELISA test was used to detect IgG in human sera of infected COVID-19 patients, and showed sensitivity and specificity of 69.6% and 88%, respectively. Recently, Villafañe et al [30], showed 100% serological concordance, using an in-house ELISA test based on recombinant RBD expressed in mammalian system, with commercial test based on the full-length spike protein (COVIDAR). However, the sensitivity of COVIDAR test, reported for IgG detection was 72% to 74% between 2 and 3 weeks from the onset of symptoms and seroconversion increased up to 90.4% after 3 weeks [31].…”

Section: Coronavirus Disease 2019 (Covid-19mentioning

confidence: 99%

Assessment of specific human antibodies against SARS-CoV-2 receptor binding domain by rapid in-house ELISA

Hussein

Ali

El-Hakim

et al. 2022

HAB

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BACKGROUND: The recently emerged SARS-CoV-2 caused a global pandemic since the last two years. The urgent need to control the spread of the virus and rapid application of the suitable health measures raised the importance of available, rapid, and accurate diagnostic approaches. OBJECTIVE: The purpose of this study is to describe a rapid in-house optimized ELISA based on the expression of the receptor binding domain (RBD) of the SARS-CoV-2 spike protein in a prokaryotic system. METHODS: We show the expression of the 30 kDa recombinant SARS-CoV-2 RBD-6×His in five different E. coli strains (at 28∘C using 0.25mM IPTG) including the expression strain E. coli BL21 (DE3) Rosetta Gami. SARS-CoV-2 rRBD-6×His protein was purified, refolded, and used as an antigen coat to assess antibody response in human sera against SARS-CoV-2 infection. RESULTS: The assessment was carried out using a total of 155 human sero-positive and negative SARS-CoV-2 antibodies. The ELISA showed 69.5% sensitivity, 88% specificity, 78.5% agreement, a positive predictive value (PPV) of 92.3%, and a negative predictive value of 56.5%. Moreover, the optical density (OD) values of positive samples significantly correlated with the commercial kit titers. CONCLUSIONS: In conclusion, specific human antibodies against SARS-CoV-2 spike protein were detected by rapid in-house ELISA in sera of human COVID-19-infected patients. The availability of this in-house ELISA protocol would be valuable for various diagnostic and epidemiological applications, particularly in developing countries. Future studies are planned for the use of the generated SARS-CoV-2 rRBD-6×His protein in vaccine development and other diagnostic applications.

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“…An increased level of IgG in blood is associated with disorders like hyperthyroidism and systemic lupus erythematosus. The detection of IgG is also important to test the presence of novel coronavirus in human body as well as to assess the efficacy of vaccines during the course of immunization 10 , 11 . The normal range of IgG in human blood range from 6 to 16 g/L.…”

Section: Introductionmentioning

confidence: 99%

L-cysteine/MoS2 modified robust surface plasmon resonance optical fiber sensor for sensing of Ferritin and IgG

Thawany

Khanna

Tiwari

et al. 2023

Sci Rep

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L-cysteine conjugated molybdenum disulphide (MoS2) nanosheets have been covalently attached to a gold coated surface plasmon resonance (SPR) optical fiber to prepare a robust and stable sensor. Owing to the multifunctionality of the deposited nanosheet conjugate, the antibodies are also covalently conjugated in the subsequent step to realize the design of a SPR optical fiber biosensor for the two important bioanalytes namely, Ferritin and Immunoglobin G (IgG). The different stages of the biosensor preparation have been characterized and verified with microscopic and spectroscopic techniques. A uniform and stable deposition of the L-cysteine/MoS2 nanosheets has allowed the biosensor to be reused for multiple times. Unlike the peeling-off of the MoS2 coatings from the gold layer reported previously in the case of physically adsorbed nanomaterial, the herein adopted strategy addresses this critical concern. It has also been possible to use the single SPR fiber for both Ferritin and IgG bioassay experiments by regenerating the sensor and immobilizing two different antibodies in separate steps. For ferritin, the biosensor has delivered a linear sensor response (SPR wavelength shifts) in the concentration range of 50–400 ng/mL, while IgG has been successfully sensed from 50 to 250 µg/mL. The limit of detection for Ferritin and IgG analysis have been estimated to be 12 ng/mL and 7.2 µg/mL, respectively. The biosensors have also been verified for their specificity for the targeted molecule only. A uniform and stable deposition of the nanomaterial conjugate, reproducibility, regeneration capacity, a good sensitivity, and the specificity can be highlighted as some of key features of the L-cysteine/MoS2 optical fiber biosensor. The system can be advocated as a useful biosensor setup for the sensitive biosensing of Ferritin and IgG.

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Development and evaluation of a low cost IgG ELISA test based in RBD protein for COVID-19

Cited by 12 publications

References 14 publications

Design and optimization of an IgG human ELISA assay reactive to recombinant RBD SARS-CoV-2 protein

Design and optimization of an IgG human ELISA assay reactive to recombinant RBD SARS-CoV-2 protein

Assessment of specific human antibodies against SARS-CoV-2 receptor binding domain by rapid in-house ELISA

L-cysteine/MoS2 modified robust surface plasmon resonance optical fiber sensor for sensing of Ferritin and IgG

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