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. 2023 Mar 27;11(4):743.
doi: 10.3390/vaccines11040743.

VLP-ELISA for the Detection of IgG Antibodies against Spike, Envelope, and Membrane Antigens of SARS-CoV-2 in Indian Population

Dilip Kumar  1 , Sourav Singha Roy  1 , Ruchir Rastogi  1 , Kajal Arora  1 , Avinash Undale  1 , Reeshu Gupta  1   2 , Nupur Mehrotra Arora  1 , Prabuddha K Kundu  1
Affiliations

VLP-ELISA for the Detection of IgG Antibodies against Spike, Envelope, and Membrane Antigens of SARS-CoV-2 in Indian Population

Dilip Kumar et al. Vaccines (Basel). .

Abstract

Background: Serological methods to conduct epidemiological survey are often directed only against the spike protein. To overcome this limitation, we have designed PRAK-03202, a virus-like particle (VLP), by inserting three antigens (Spike, envelope and membrane) of SARS-CoV-2 into a highly characterized S. cerevisiae-based D-Crypt™ platform.

Methods: Dot blot analysis was performed to confirm the presence of S, E, and M proteins in PRAK-03202. The number of particles in PRAK-03202 was measured using nanoparticle tracking analysis (NTA). The sensitivity of VLP-ELISA was evaluated in 100 COVID positive. PRAK-03202 was produced at a 5 L scale using fed-batch fermentation.

Results: Dot blot confirmed the presence of S, E, and M proteins in PRAK-03202. The number of particles in PRAK-03202 was 1.21 × 109 mL-1. In samples collected >14 days after symptom onset, the sensitivity, specificity, and accuracy of VLP-ELISA were 96%. We did not observe any significant differences in sensitivity, specificity, and accuracy when post-COVID-19 samples were used as negative controls compared to pre-COVID-samples. At a scale of 5 L, the total yield of PRAK-03202 was 100-120 mg/L.

Conclusion: In conclusion, we have successfully developed an in-house VLP-ELISA to detect IgG antibodies against three antigens of SARS-CoV-2 as a simple and affordable alternative test.

Keywords: COVID-19; ELISA; immunoglobulin G; nanoparticle tracking analysis; virus-like particles.

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Conflict of interest statement

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
(A) Dot blot analysis for co-expression of spike, envelope, and membrane proteins in the S. cerevisiae-based D-Crypt™ platform using antigen-specific antibodies S: spike protein, E: envelope protein, M: membrane protein. (B) Nanoparticle tracking analysis with 0.25 (left panel) and 0.5 mg (right panel) of PRAK-03202. (C) Optimum time to obtain maximum production of PRAK-03202 in the fermenter. The fermenter was harvested at 24, 48, and 72 h to determine the maximum OD of the culture.
Figure 2
Figure 2
Multiple sequence alignment of BQ.1.1 and Wuhan variant of SARS-CoV-2. Fully conserved residues, insertion/deletion of amino acids, and non-conserved amino acids are indicated by *, ---, and a gap, respectively.
Figure 2
Figure 2
Multiple sequence alignment of BQ.1.1 and Wuhan variant of SARS-CoV-2. Fully conserved residues, insertion/deletion of amino acids, and non-conserved amino acids are indicated by *, ---, and a gap, respectively.
Figure 2
Figure 2
Multiple sequence alignment of BQ.1.1 and Wuhan variant of SARS-CoV-2. Fully conserved residues, insertion/deletion of amino acids, and non-conserved amino acids are indicated by *, ---, and a gap, respectively.
Figure 2
Figure 2
Multiple sequence alignment of BQ.1.1 and Wuhan variant of SARS-CoV-2. Fully conserved residues, insertion/deletion of amino acids, and non-conserved amino acids are indicated by *, ---, and a gap, respectively.
Figure 3
Figure 3
(A) IgG response against PRAK-03202 in COVID-19-negative and -positive convalescent samples. ---- indicates cut-off values. (B) ROC curve for the VLP-ELISA using optical density values.
Figure 4
Figure 4
IgG response against PRAK-03202 in (A,B) COVID-19-positive convalescent samples at the indicated periods of symptoms onset and negative samples from (A) pre-COVID-19 and (B) post-COVID-19 era. ---- indicates the cut-off values.
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