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. 2023 Jul 6:11:1202126.
doi: 10.3389/fbioe.2023.1202126. eCollection 2023.

New detection method of SARS-CoV-2 antibodies toward a point-of-care biosensor

Janikua Nelson-Mora  1 , Diana Rubio  1 , Amairani Ventura-Martínez  1 , Luis A González  1 , Diana Del-Rio  1   2 , Yuli Aranda-López  1 , Edgar Jiménez-Díaz  2   3 , Diego Zamarrón-Hernández  1 , Diana G Ríos-López  1 , Stephanie Aguirre  1 , Yasab Ruiz-Hernandez  1 , Aarón Cruz-Ramírez  1 , Jonás S Barjau  1 , Miguel A Jáurez  1 , Jehú Lopez-Aparicio  1 , Andrea Campa-Higareda  1 , Tatiana Fiordelisio  1   2   3
Affiliations

New detection method of SARS-CoV-2 antibodies toward a point-of-care biosensor

Janikua Nelson-Mora et al. Front Bioeng Biotechnol. .

Abstract

The outbreak of COVID-19, a disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, is regarded as the most severe of the documented coronavirus pandemics. The measurement and monitoring of SARS-CoV-2 antibody levels by serological tests are relevant for a better epidemiological and clinical understanding of COVID-19. The aim of this work was to design a method called the SARS-CoV-2 antibody detection method (SARS-CoV-2 AbDM) for fluorescence immunodetection of anti-SARS-CoV-2 IgG and IgM on both plate and microfluidic chip. For this purpose, a system with magnetic beads that immobilize the antigen (S protein and RBD) on its surface was used to determine the presence and quantity of antibodies in a sample in a single reaction. The SARS-CoV-2 AbDM led to several advantages in the performance of the tests, such as reduced cost, possibility of performing isolated or multiple samples, potential of multiplex detection, and capacity to detect whole blood samples without losing resolution. In addition, due to the microfluidic chip in conjunction with the motorized actuated platform, the time, sample quantity, and operator intervention during the process were reduced. All these advantages suggest that the SARS-CoV-2 AbDM has the potential to be developed as a PoC that can be used as a tool for seroprevalence monitoring, allowing a better understanding of the epidemiological and clinical characteristics of COVID-19 and contributing to more effective and ethical decision-making in strategies to fight against the COVID-19 pandemic.

Keywords: COVID-19; fluorescence; immunodetection; magnetic beads; microfluidic chip.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Microfluidic chip for SARS-CoV-2 AbDM. (A) Schema of the sequence of microchannel and the contents of each well. (B) Photograph of the automated platform used to move the magnetic beads in the microfluidic chip. (C) Sealing of the microfluidic chip. The microstructured PMMA is coated in uncured PDMS and then sealed with a coverslip.
FIGURE 2
FIGURE 2
Optimization of the SARS-CoV-2 AbDM for IgG-RBD detection. (A) Plot of the fluorescence intensity obtained for different concentrations of RBD, used in the functionalization of the beads (360K beads), and (B) for different concentrations of αHIgG-647. (C) Comparative plot of the fluorescence and absorbance measurement obtained for IgG-RBD with the SARS-CoV-2 AbDM and with ELISA, respectively. Plotted (●) is the result of SARS-CoV-2 AbDM and (◻) is the values obtained by ELISA. In (B) and (C), Δ represents the fluorescence value of the control beads functionalized without RBD (woRBD). In all plots, symbols represent the mean of three replicates ± SD. Statistically significant differences are shown with **** (p < 0.0001) for SARS-CoV-2 AbDM and with #### (p < 0.0001) for ELISA. (D–F) Representative confocal microscopy images of (A). Scales: 50 μm.
FIGURE 3
FIGURE 3
Optimization of the SARS-CoV-2 AbDM for IgG-S detection. (A) Plot of the fluorescence intensity obtained for beads (240K beads) functionalized with different concentrations of S protein and (B) for different concentrations of αHIgG-647. (C) Comparison of the fluorescence and absorbance measured for IgG-S using the SARS-CoV-2 AbDM and ELISA, respectively. Plot shows (●) the SARS-CoV-2 AbDM results and (◻) the ELISA values. In (B) and (C), Δ represents the fluorescence intensity of the control beads functionalized without S protein (woS). In all plots, symbols represent the mean of three replicates ± SD. Statistically significant differences are shown with ** (p < 0.01) and **** (p < 0.0001) for SARS-CoV-2 AbDM and with ####, (p < 0.0001) for ELISA. (D–F) Representative confocal microscopy images of (A). Scales: 50 μm.
FIGURE 4
FIGURE 4
Serum antibody detection comparison between SARS-CoV-2 AbDM and ELISA. Samples were analyzed by both methods to detect (A–C) IgG-RBD (n = 50) and (D–F) IgG-S (n = 31). (A and D) ELISA titer versus SARS-CoV-2 AbDM normalized fluorescence intensity. The mean (●) of the SARS-CoV-2 AbDM result for each sample and the mean and deviation of each ELISA titer population are shown. woRBD and woS correspond to control beads functionalized without antigen. (B and E) Receiver operating characteristic (ROC) curve for SARS-CoV-2 AbDM, with an area under the curve of 0.84 for IgG-RBD and 1 for IgG-S. (C and F) SARS-CoV-2 AbDM titer versus ELISA titer. Cut-off points of the normalized fluorescence intensity were determined for each titer using ROC analysis. The size of the bubble is proportional to the number of incidences (1–10) in each range. All negative cases were plotted with the value of 10, as the scale is logarithmic.
FIGURE 5
FIGURE 5
Detection of (A) IgG-RBD and (B) IgG-S in serum and whole blood using SARS-CoV-2 AbDM in ELISA-titrated samples. Mean of three replicates ± SD of (●) serum and (◻) whole blood fluorescence intensity values are plotted. woRBD and woS correspond to control beads functionalized without antigen and reacted with the highest titered sample. Statistically significant differences are shown with **** (p < 0.0001) for serum and with #### (p < 0.0001) for whole blood.
FIGURE 6
FIGURE 6
Co-detection of IgG and IgM in serum samples with SARS-CoV-2 AbDM, using (A) RBD and (B) S protein as antigen. Mean of three replicates ± SD of (●) IgG (αHIgG-647) and (◻) IgM (αHIgM-488) fluorescence intensity values are plotted. Two serum samples previously analyzed by ELISA were tested: a serum sample positive only for IgG (+IgG/-IgM) and a serum sample positive for both antibodies (+IgG/+IgM); woRBD and woS correspond to control beads functionalized without antigen and reacted with the +IgG/+IgM sample. Statistically significant differences by ANOVA are shown for IgG **** (p < 0.0001) and for IgM ## (p < 0.001) and # (p < 0.01). (C–F) Representative confocal microscopy images of IgG and IgM detection with both antigens. Scales: 50 μm.
FIGURE 7
FIGURE 7
Optimization of IgG-S detection with SARS-CoV-2 AbDM for on-chip microfluidic implementation. (A) Fluorescence intensity plot for different secondary antibody (αHIgG-647) concentrations on-chip. (B) Comparison of fluorescence and absorbance measurements for IgG-S with SARS-CoV-2 AbDM on-chip and with ELISA, respectively. The SARS-CoV-2 AbDM results are shown with (●), and the ELISA values are shown with (◻). In (A) and (B), (Δ) represents the fluorescence intensity of the control beads functionalized without S protein (woS). (C) Determination of serum sample dilution ratio. Fluorescence intensity of samples with different ELISA titers (negative; 50; 8100) are shown with (●) for 1:6, with (◻) for 1:3, and with (▲) for 2:3. (D) ELISA titer versus normalized fluorescence intensity of SARS-CoV-2 AbDM on-chip. The mean (●) of the SARS-CoV-2 AbDM result on chip for each sample (n = 21) and the mean and deviation of each ELISA titer population are shown. (E) ROC curve for SARS-CoV-2 AbDM on chip, with an area under the curve of 0.94 for IgG-S. Symbols represent the mean of three (A–C) or two (D) replicates ± SD. Statistically significant differences are shown with *** (p < 0.001) for SARS-CoV-2 AbDM and with #### (p < 0.0001) for ELISA.
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References

    1. Abdulamir A. S., Hafidh R. R. (2020). The possible immunological pathways for the variable immunopathogenesis of COVID—19 infections among healthy adults, elderly and children. Electron. J. Gen. Med. 17, em202. 10.29333/ejgm/7850 - DOI
    1. Amanat F., Stadlbauer D., Strohmeier S., Nguyen T. H. O., Chromikova V., McMahon M., et al. (2020). A serological assay to detect SARS-CoV-2 seroconversion in humans. Nat. Med. 26, 1033–1036. 10.1038/s41591-020-0913-5 - DOI - PMC - PubMed
    1. Azkur A. K., Akdis M., Azkur D., Sokolowska M., van de Veen W., Brüggen M.-C., et al. (2020). Immune response to SARS-CoV-2 and mechanisms of immunopathological changes in COVID-19. Allergy 75, 1564–1581. 10.1111/all.14364 - DOI - PMC - PubMed
    1. Bachmann M. F., Mohsen M. O., Zha L., Vogel M., Speiser D. E. (2021). SARS-CoV-2 structural features may explain limited neutralizing-antibody responses. NPJ Vaccines 6, 2. 10.1038/s41541-020-00264-6 - DOI - PMC - PubMed
    1. Bong C.-L., Brasher C., Chikumba E., McDougall R., Mellin-Olsen J., Enright A. (2020). The COVID-19 pandemic: Effects on low- and middle-income countries. Anesth. Analg. 131, 86–92. 10.1213/ane.0000000000004846 - DOI - PMC - PubMed

Grants and funding

This work was funded by UNAM-PAPIIT BV-200820, CONACYT CY-313005, CONACYT CY-315805, Liomont Labs, Kaluz Foundation, Casa Cordoba, Sertull Foundation, Roberto Hernandez R Foundation, and Ph.D. Armando Jinich Ripstein.