Non-tumorigenic MDCK cell line for propagating viruses
The present invention provides novel MDCK-derived adherent non-tumorigenic cell lines that can be grown in the presence or absence of serum. The cell lines of the present invention are useful for the production of vaccine material (e.g., viruses). More specifically, the cell lines of the present invention are useful for the production of influenza viruses in general and ca/ts influenza viruses in particular. The invention further provides methods and media formulations for the adaptation and cultivation of MDCK cells such that they remain non-tumorigenic. Additionally, the present invention provides methods for the production of vaccine material (e.g., influenza virus) in the novel cell lines of the invention.
1. A process for the production of influenza viruses comprising:
a) infecting adherent and non-tumorigenic Madin Darby Canine Kidney (MDCK) cells with influenza viruses, which cells are adapted to grow and cultivated in serum-free medium comprising Taub's medium, lipids and wheat hydrolysate, which Taub's medium comprises a 50:50 mixture of DMEM and Ham's F12, 4.5 grams per liter glucose, 4 millimolar L-glutamine, 5 micrograms per milliliter insulin, 5 micrograms per milliliter transferrin, 5 picomolar triiodothyronine, 50 nanomolar hydrocortisone, 25 nanograms per milliliter prostaglandin E1 and 10 nanomolar sodium selenite;
b) incubating the cells; and
c) isolating the influenza viruses from the cells.
2. The process of claim 1 , wherein the serum-free medium is MediV SF103.
3. The process of claim 1 , wherein the serum-free medium further comprises epidermal growth factor (EGF).
4. The process of claim 3 , wherein the serum-free medium is MediV SF104.
5. The process of claim 3 , wherein the serum-free medium further comprises tropolone and lacks transferrin.
6. The process of claim 5 , wherein the serum-free medium is MediV SF105.
7. The process of claim 1 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
8. The process of claim 2 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
9. The process of claim 3 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
10. The process of claim 4 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
11. The process of claim 5 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
12. The process of claim 6 , wherein the MDCK cells are from the cell line MDCK-S, deposited as ATCC Accession No. PTA-6500; are from the cell line deposited as ATCC Accession No. CCL34; or are from the cell line MDCK-SF103, deposited as ATCC Accession No. PTA-6503.
13. The process of claim 1 , wherein a protease is added before, during, or after step (a).
14. The process of claim 2 , wherein a protease is added before, during, or after step (a).
15. The process of claim 3 , wherein a protease is added before, during, or after step (a).
16. The process of claim 4 , wherein a protease is added before, during, or after step (a).
17. The process of claim 5 , wherein a protease is added before, during, or after step (a).
18. The process of claim 6 , wherein a protease is added before, during, or after step (a).
19. The process of claim 1 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
20. The process of claim 2 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
21. The process of claim 3 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
22. The process of claim 4 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
23. The process of claim 5 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
24. The process of claim 6 , wherein the influenza virus is temperature-sensitive, and/or cold-adapted, and/or attenuated.
25. The process of claim 1 , wherein the MDCK cells are cultivated on microcarriers.