Coronavirus proteins and antigens
Disclosed herein are embodiments of a method for collecting, extracting or eluting proteins and antigens from cells infected with coronavirus. The coronavirus may be a porcine coronavirus, such as porcine epidemic diarrhea virus (PEDV) or porcine delta coronavirus (PDCoV). Also disclosed are embodiments of a composition comprising the coronavirus proteins and antigens, and embodiments of a method of using such a composition. Applications for the composition include, but are not limited to, use in the preparation of antibodies against the proteins and antigens, use as reference markers for coronavirus proteins, and/or use in an immunogenic composition, such as in a vaccine composition.
1. A method, comprising:
isolating porcine epidemic diarrhea virus (PEDV)-infected cells away from cell-free PEDV viral particles in a population of cells in a culture medium;
extracting or eluting PEDV proteins and antigens from the isolated PEDV-infected cells with a detergent-containing solution to form a first solution comprising detergent and isolated PEDV proteins and antigens; and
adding an adjuvant to the first solution to form a second solution comprising the isolated PEDV proteins and antigens, the detergent and the adjuvant.
2. The method of claim 1 , further comprising allowing the population of cells to incubate for a time period sufficient to result in one or more replicated PEDV viral particles being released into the culture medium.
3. The method of claim 1 , wherein the detergent-containing solution comprises a non-ionic detergent.
4. The method of claim 3 , wherein the non-ionic detergent is poly(ethylene glycol) p-isooctyl-phenyl ether, octylphenoxypolyethoxyethanol (Nonidet P-40), 4-(1,1,3,3-tetramethylbutyl)phenyl-polyethylene glycol (Triton X-100), or a combination thereof.
5. The method of claim 3 , wherein the detergent-containing solution comprises a detergent at a concentration of greater than zero to 5% in solution.
6. The method of claim 3 , wherein the detergent-containing solution comprises a chelating agent at a concentration of greater than zero to 0.5M in solution.
7. The method of claim 1 , wherein extracting or eluting PEDV proteins and antigens comprises extracting or eluting PEDV proteins and antigens for about 0.1 to about 15 hours.
8. The method of claim 7 , wherein extracting or eluting PEDV proteins and antigens is performed at a temperature of from 0° C. to 25° C.
9. The method of claim 7 , wherein extracting or eluting PEDV proteins and antigens comprises extracting or eluting PEDV proteins and antigens for from 0.1 hours to 15 hours at 4° C.
10. The method of claim 1 , wherein the proteins and antigens include PEDV envelope proteins.
11. The method of claim 1 , wherein the first solution of isolated PEDV proteins and antigens has a concentration of PEDV proteins and antigens greater than a concentration of PEDV proteins and antigens in the culture medium that contained the population of infected cells.
12. The method of claim 1 , further comprising adding an excipient to the first solution or the second solution.
13. A method, comprising:
isolating PEDV-infected cells away from cell-free PEDV viral particles in a population of cells in a culture medium;
allowing the population of cells to incubate for a time period sufficient to result in one or more replicated PEDV viral particles being released into the culture medium;
extracting or eluting PEDV proteins and antigens from the isolated PEDV-infected cells with a detergent-containing solution to form a solution of isolated PEDV proteins and antigens; and
adding an adjuvant to the solution of isolated PEDV proteins and antigens, to form an immunogenic composition comprising PEDV proteins and antigens, cell fragments, the detergent and the adjuvant.
14. A composition comprising isolated PEDV proteins and antigens from claim 1 , an adjuvant, and a detergent made by the method of claim 1 .
15. The composition of claim 14 , comprising PEDV proteins and antigens, cell fragments, the detergent and the adjuvant.
16. A method of vaccinating a subject, the method comprising administering to the subject the second solution from claim 1 or a composition comprising the second solution of isolated PEDV proteins and antigens from claim 1 .
17. The method of claim 16 , wherein the subject is a porcine subject.
18. A method of generating an immune response in a porcine subject, the method comprising administering to the subject the composition of claim 15 .
19. The method of claim 6 , wherein the chelating agent is ethylenediaminetetraacetic acid.
20. The composition of claim 14 , wherein the composition comprises the isolated PEDV proteins and antigens, the adjuvant, the detergent, and ethylenediaminetetraacetic acid.