ELISpot Serological Assays for Coronavirus Research
Highly Sensitive and Specific B Cell ELISpot Assays for SARS-CoV-2 IgG and IgM Antibodies.
B Cell ELISpot Assays to Measure the Immune Response to Infection
The antigen-down B cell ELISpot assay is a powerful 96-well assay to study the extent of the immune response to viral infections. The larger the number of spots left by B cells, the stronger the antibody-producing capacity of the immune system. The B cell ELISpot is built with highly sensitive and specific antibodies. When paired with pure and specific SARS-CoV-2 proteins, it becomes a robust and highly sensitive serological assay for SARS-CoV-2 infection.
Investigators at the University of Minnesota utilized R&D Systems ELISpot plates pre-coated with SARS-CoV-2 recombinant proteins to assess IgM and IgG secretion by B cells collected from volunteers (Figure 1).
A
B
C
Figure 1 B Cells from a volunteer (1 month post-SARS-CoV-2 infection) were cultured in plates pre-coated with 15 ug/ml of SARS-CoV-2 Spike RBD His-Tag Protein (A), SARS-CoV-2 Spike (Active Trimer) His Protein (B) and SARS-CoV-2 Spike (GCN4-IZ) His Protein (C) at a density of 50,000 cells per well. Reactive spots indicate IgG secretion from cultured B cells. The larger the spot the more robust the IgG secretion. Cells from uninfected volunteers were not reactive (Data not shown).
Protocol: B-Cell ELISpot as a SARS-CoV-2 Single Color IgG or IgM Serology Test
R&D Systems IgG and IgM Antibodies are Specific
R&D Systems manufactures highly specific IgG and IgM antibodies. Direct ELISAs are commonly used to evaluate antibody affinity. Figures 2 and 3 demonstrate the specificity of the anti-human IgG (hIG) and anti-human IgM (hIgM) antibodies antibody provided in our B-cell ELISpot assay have a high affinity for hIgG and hIgM, respectively, with limited cross reactivity to other classes of immunoglobulins.
Figure 2. Data from a direct ELISA indicates that the R&D Systems anti-human IgG antibody included in R&D Systems B cell ELISpot assays is specific for hIgG 1-4 with minimal cross reactivity to hIgA, hIgD, hIgE or hIgM.
Figure 3. Data from a direct ELISA indicates that the R&D Systems anti-human IgM antibody included in R&D Systems B cell ELISpot assays is specific for hIgM with no cross-reactivity to hIgA, hIgG, hIgG Fc only.
High Quality Recombinant Coronavirus Proteins
The data described in figure 1 was generated using high-quality R&D Systems recombinant coronavirus proteins as antigens. The data examples shown below demonstrate the purity (Figure 4) and lot-to-lot consistency (Figure 5) of recombinant SARS-CoV-2 Spike proteins. Download this flyer for a more detailed description of analyses that R&D Systems scientists routinely perform to characterize our antibodies and recombinant proteins prior to their commercial release.
Figure 4. The purity of Recombinant SARS-CoV-2 Spike RBD His-tag Protein was assessed by SDS-PAGE under reducing (R) and non-reducing (NR) conditions and visualized by Coomassie® Blue staining, showing bands at the expected molecular weight range of 32-38 kDa.
Figure 5. Four independent lots of Recombinant Human ACE-2 His-tag Protein were tested for their ability to bind to Recombinant SARSCoV-2 Spike RBD Fc Chimera Protein in a functional ELISA. The data demonstrates high lot-to-lot consistency across these four lots.
Table 1: Available recombinant coronavirus proteins. The first three proteins were used as antigens in figure 1.
| Catalog | Recombinant Protein |
|---|---|
| 10500-CV | SARS-CoV-2 Spike RBD His-Tag Protein |
| 10549-CV | SARS-CoV-2 Spike (Active Trimer) His Protein |
| 10561-CV | SARS-CoV-2 Spike (GCN4-IZ) His Protein |
| 10522-CV | SARS-CoV-2 Spike S1 |
| 10523-CV | SARS-CoV-2 Spike RBD |
| 10474-CV | SARS-CoV-2 Nucleocapsid |
| 10521-CV | MERS-CoV Nucleocapsid |
| 10621-CV | MERS-CoV Spike RBD |
| 10600-CV | HCoV-HKU1 Spike RBD |
| 10605-CV | HCoV-NL63 Spike RBD Protein |
| 10612-CV | HCoV-229E Spike RBD Protein |
Don’t see the specific protein you need? We have a large selection of recombinant coronavirus proteins.
Conclusions
R&D Systems B cell ELISpot Assays can be utilized as a sensitive and specific, economical coronavirus serological research tool. Together, R&D Systems B cell ELISpot and recombinant coronavirus proteins can be used in your research to study the immune response to natural SARS-CoV-2 infection or vaccine candidates. When used with high-quality R&D Systems recombinant proteins, you can expect reliable and consistent data over the long term.
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