Ha-CoV-2

A new proprietary hybrid alphavirus-SARS-CoV-2 pseudovirion

Ha-CoV-2 is a newly developed hybrid SARS-CoV-2 virus-like particle (VLP) that encapsulates an alphavirus-derived RNA genome for rapid reporter expression (luciferase or GFP) in target cells (Hetrick et al., 2020)(patent pending). Different from commonly used S protein pseudotyped lenti- or vesicular stomatitis virus (VSV)-psedoviruses, Ha-CoV-2 is assembled with all 4 structural proteins (S, M, N, and E) of SARS-CoV-2, and contains a reporter genome derived from an alphavirus-based vector for rapid (6 hours) and robust expression of reporter genes. Ha-CoV-2 represents a major technology advancement in the development of SARS-CoV-2 pseudoviruses, and serves as platforms for rapid and robust quantification of neutralizing antibodies, viral mutants, and antiviral drugs (Dabbagh et al., 2021; He et al., 2021).

Virongy offers pre-assembled Ha-CoV-2 reporter pseudovirions. We also help customers to perform antiviral drug screening and quantification of neutralizing antibodies. For more information, please contact us by email: info@virongy.com

Fig. 1. Schematic of the assembly of Ha-CoV-2 particles.

 

Major advances

  • Faster speed – As fast as 6 hours for the detection of reporter expression for Ha-CoV-2 versus 2-3 days for S protein pseudotyped lentivirus.
  • More robust reporter signal - Ha-CoV-2 takes advantage of the rapid and robust gene expression capacity of alphavirus vector for reporter expression.
  • High fidelity in particle structure – Ha-CoV-2 contains all 4 SARS-CoV-2 structural proteins (S, M, E, and N), but no structural proteins from other viruses. In contrast, lenti- and VSV- pseudoviruses contain only the S protein from SARAS-CoV-2, and multiple structural proteins from other viruses, such as HIV-1 Gag and Pol.

 

 

 

 

 

 

Fig. 2. Comparison of Ha-CoV-2 with S protein pseudotyped lentivirus in a time course of infection and reporter expression.

HEK293T(ACE2/TMPRESS2) cells were used as the target cell.

 

Applications

  • Screening and quantification of anti-SARS-CoV-2 neutralizing antibodies
  • Anti-SARS-CoV-2 drug screening
  • Quantification of SARS-CoV-2 viral mutants
  • Identification of host co-factors and restriction factors of SARS-CoV-2

Fig. 3. Neutralizing antibody activity measured with Ha-CoV-2(Luc).

50 μl of Ha-CoV-2(Luc) (Cat# HaCoV2Luc-01) was incubated with serially diluted anti-serum for 1 hour at 37°C. The virus-antibody complex was used to infect 40,000 HEK293T(ACE2/TMPRSS2) target cells in a 96-well plate. Luciferase assay was performed at 12 hours post infection.

 

Table 1. Ha-CoV-2 reporter pseudovirion

Catalog No. Products Price1
HaCoV2Luc-01 5 x 200 μl of Ha-CoV-2 (Luc) particles, Luciferase reporter. $350.00
HaCoV2Luc-02 10 x 200 μl of Ha-CoV-2 (Luc) particles, Luciferase reporter. $680.00
HaCoV2Luc-03 50 x 200 μl of Ha-CoV-2 (Luc) particles, Luciferase reporter. $3200.00
HaCoV2Luc-04 100 x 200 μl of Ha-CoV-2(Luc) particles, Luciferase reporter. $6200.00
HaCoV2Luc-10C 5 x 100 μl of Ha-CoV-2(Luc) particles (concentrated), Luciferase reporter. $1,900.00
HaCoV2GFP-01 5 x 200 μl of Ha-CoV-2(GFP) particles, GFP reporter. $350.00
HaCoV2GFP-10C 5 x 100 μl of Ha-CoV-2(GFP) particles (concentrated), GFP reporter. $1,900.00
1Academic and government price. Others inquire.
Technical Support Email: info@virongy.com

 

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Suggested protocol

(For the infection of target cells in 12-well plate. For more protocol details in other formats, such as in 96-well plate, please contact info@virongy.com)

  • Count target cells, g. HEK293T(ACE2/TMPRSS2), to be infected, and seed 1 x 105 cells per well in 0.5 ml culture medium in each well of 12-well plate. Culture cells overnight. Note: Cell viability should be ≥ 80%.
  • Before infection, remove medium, add 300 μl fresh medium.
  • Add 100 μl Ha-CoV-2 virus, and infect for 2-6 hours. *
  • Wash cells by adding 1 ml fresh medium.
  • After washing, add 1 ml fresh complete medium (DMEM + 10% FBS).
  • Culture cells for 12-18 hours to read signals (e.g. luciferase assay).

* if performing neutralizing antibody assays, incubate virus with antibodies at 37°C for 1 hour, then add the virus-antibody complex to cells for infection.

Ha-CoV-2  are intended for Research Use Only and are not for diagnostic or therapeutic purposes or uses in humans or animals.

 

References

Dabbagh, D., He, S., Hetrick, B., Chilin, L., Andalibi, A., and Wu, Y. (2021). Identification of the SHREK family of proteins as broad-spectrum host antiviral factors. bioRxiv, doi: https://doi.org/10.1101/2021.02.02.429469

He, S., Waheed, A.A., Hetrick, B., Dabbagh, D., Akhrymuk, I.V., Kehn-Hall, K., Freed, E.O., and Wu, Y. (2021). PSGL-1 Inhibits the Incorporation of SARS-CoV and SARS-CoV-2 Spike Glycoproteins into Pseudovirions and Impairs Pseudovirus Attachment and Infectivity. Viruses 13, 46.

Hetrick, B., He, S., Chilin, L.D., Dabbagh, D., Alem, F., Narayanan, A., Luchini, A., Li, T., Liu, X., Liotta, L., et al. (2020). Development of a novel hybrid alphavirus-SARS-CoV-2 particle for rapid in vitro screening and quantification of neutralization antibodies, antiviral drugs, and viral mutations. bioRxiv, doi: https://doi.org/10.1101/2020.12.22.423965