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June 12, 2020
NLSinsight

Q+A with Mabtech's Head of Research & Development, Bartek Makower- COVID-19 Special.

Q+A with Mabtech's Head of Research & Development, Bartek Makower- COVID-19 Special.

-
NLSinsight
-
June 12, 2020



Bartek Makower, PhD
Head of Research and Development


Mabtech are reknown for supplying immune assays and instruments to study immune responses, especially in vaccine development, I can imagine this has been an extremely busy period. Are there specific products you supply that have been in high demand?

We have seen a big increase in the purchases of IFN-gamma ELISpot kits for human, mouse as well as non-human primate samples. We think this has to do with IFN-gamma ELISpot being such a high-throughput and robust assay, allowing pharma companies to easily screen their vaccine candidates for immunogenicity and efficacy. We have also seen a surge in demand for IL-6 ELISA kits, tentatively due to IL-6 being shown to correlate with disease severity in COVID-19 patients.


Across the frontpage of Mabtechs websites it states ‘Don’t forget about T cells and memory B cells when studying COVID-19’ – tell us more about this and why you think its so important.

Looking at the SARS-CoV-2 literature it is quite clear that that 10-20% of infected individuals develop no or very weak antibody responses when measured by ELISA. Without CD4+ T-cell responses, you will not get IgG or IgA responses, thus these cells are essential to study. Cytotoxic CD8+ T-cells kill virus infected cells and the frequency of such cells may be correlated to severity of disease. A recent publication compared five different methods to measure antibodies (three ELISA methods, one immunofluorescence method based on infected cells and one virus neutralization method) to the B cell FluoroSpot assay. The B cell FluoroSpot detected antibody secreting cells in 100% of the patients while the antibody methods only detected responses in 75-80% of the patients. We thus believe that looking at the full picture, i.e. antibodies in plasma, T cells and B cells is essential for several reasons:

i. How many have been infected in the population? ELISA alone will not detect all previously infected individuals.

ii. Mapping T-cell responses in essential in vaccine design. Literature shows that SARS-CoV-2 infection induces a Th1 response. A vaccine which induces a Th2 response might enhance disease.

iii. And finally, mapping of the natural immune response induced by SARS-CoV-2 is also essential for several reasons:

- It will aid in vaccine design.
- If patients are followed overtime, we can answer the question about immunity. Is it protective and how long does it last?
- We might find immune response patterns which predict if a patient will get mild or severe disease.


What methods that Mabtech use can detect Memory B Cells for SARS-CoV-2?

We use reversed B cell ELISpot or FluoroSpot assays. These methods are used to detect antibody secreting cells (ASC) as early as 3-5 days after infection and memory B cells several years after infection. In order to get reliable ELISA responses one has to wait 19 days after primary infection and we don’t know how fast the response wanes. Analysis of B-cell frequencies is more sensitive and gives a much longer window of analysis.

We have chosen the spike receptor binding domain (RBD) as a target for our assays. This is because literature indicates that cross-reactive antibodies induced by prior infections by the four circulating coronaviruses are present in a high percentage of the population. According to literature the cross-reactive T-cell and antibody responses are directed to the spike S2 domain, N, M and other viral proteins, but not to S1-RBD.


You have a specific SARS-CoV-2 S2 N defined peptide pool as well as an S1 scanning pool that are now available. Tell us more about this.

a. We chose S1 as target for T-cell analysis based on the observed T-cell cross-reactivity to the other viral proteins. The S1 pool is a scanning pool of 15mers overlapping by 11. This means that it covers all possible epitopes and HLA types. As a complement to this pool we designed the S2 N pool based on mapped SARS-CoV-1 epitopes which have 100% homology to SARS-CoV-2. Together these pools give interesting information about the T-cell response in a patient. For example, if a patient has T cells specific for S2 N but no T cells to S1, this might indicate an anamnestic response due to prior exposure to a different corona virus, but not a response to SARS-CoV-2.

These peptide pools have shown to be very useful in FluoroSpot and ELISpot assays for studying T cells secreting IFN-gamma and/or IL-2. Especielly IL-2 has proven an interesting analyte, as SARS-CoV-2 specific T-cells seem to secrete more IL-2 per cell compared to other viruses like CMV, EBV and Influenza

Mabtech AB Contact Information | Biocompare.com

Last updated:
June 12, 2020

NLSinsight

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Get in touch with us!
Visit us
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Bartek Makower, PhD
Head of Research and Development


Mabtech are reknown for supplying immune assays and instruments to study immune responses, especially in vaccine development, I can imagine this has been an extremely busy period. Are there specific products you supply that have been in high demand?

We have seen a big increase in the purchases of IFN-gamma ELISpot kits for human, mouse as well as non-human primate samples. We think this has to do with IFN-gamma ELISpot being such a high-throughput and robust assay, allowing pharma companies to easily screen their vaccine candidates for immunogenicity and efficacy. We have also seen a surge in demand for IL-6 ELISA kits, tentatively due to IL-6 being shown to correlate with disease severity in COVID-19 patients.


Across the frontpage of Mabtechs websites it states ‘Don’t forget about T cells and memory B cells when studying COVID-19’ – tell us more about this and why you think its so important.

Looking at the SARS-CoV-2 literature it is quite clear that that 10-20% of infected individuals develop no or very weak antibody responses when measured by ELISA. Without CD4+ T-cell responses, you will not get IgG or IgA responses, thus these cells are essential to study. Cytotoxic CD8+ T-cells kill virus infected cells and the frequency of such cells may be correlated to severity of disease. A recent publication compared five different methods to measure antibodies (three ELISA methods, one immunofluorescence method based on infected cells and one virus neutralization method) to the B cell FluoroSpot assay. The B cell FluoroSpot detected antibody secreting cells in 100% of the patients while the antibody methods only detected responses in 75-80% of the patients. We thus believe that looking at the full picture, i.e. antibodies in plasma, T cells and B cells is essential for several reasons:

i. How many have been infected in the population? ELISA alone will not detect all previously infected individuals.

ii. Mapping T-cell responses in essential in vaccine design. Literature shows that SARS-CoV-2 infection induces a Th1 response. A vaccine which induces a Th2 response might enhance disease.

iii. And finally, mapping of the natural immune response induced by SARS-CoV-2 is also essential for several reasons:

- It will aid in vaccine design.
- If patients are followed overtime, we can answer the question about immunity. Is it protective and how long does it last?
- We might find immune response patterns which predict if a patient will get mild or severe disease.


What methods that Mabtech use can detect Memory B Cells for SARS-CoV-2?

We use reversed B cell ELISpot or FluoroSpot assays. These methods are used to detect antibody secreting cells (ASC) as early as 3-5 days after infection and memory B cells several years after infection. In order to get reliable ELISA responses one has to wait 19 days after primary infection and we don’t know how fast the response wanes. Analysis of B-cell frequencies is more sensitive and gives a much longer window of analysis.

We have chosen the spike receptor binding domain (RBD) as a target for our assays. This is because literature indicates that cross-reactive antibodies induced by prior infections by the four circulating coronaviruses are present in a high percentage of the population. According to literature the cross-reactive T-cell and antibody responses are directed to the spike S2 domain, N, M and other viral proteins, but not to S1-RBD.


You have a specific SARS-CoV-2 S2 N defined peptide pool as well as an S1 scanning pool that are now available. Tell us more about this.

a. We chose S1 as target for T-cell analysis based on the observed T-cell cross-reactivity to the other viral proteins. The S1 pool is a scanning pool of 15mers overlapping by 11. This means that it covers all possible epitopes and HLA types. As a complement to this pool we designed the S2 N pool based on mapped SARS-CoV-1 epitopes which have 100% homology to SARS-CoV-2. Together these pools give interesting information about the T-cell response in a patient. For example, if a patient has T cells specific for S2 N but no T cells to S1, this might indicate an anamnestic response due to prior exposure to a different corona virus, but not a response to SARS-CoV-2.

These peptide pools have shown to be very useful in FluoroSpot and ELISpot assays for studying T cells secreting IFN-gamma and/or IL-2. Especielly IL-2 has proven an interesting analyte, as SARS-CoV-2 specific T-cells seem to secrete more IL-2 per cell compared to other viruses like CMV, EBV and Influenza

Mabtech AB Contact Information | Biocompare.com

Last updated:
June 12, 2020

NLSinsight

Click here to read more about us!
Get in touch with us!
Visit us
Send us mail