Cells for detection of enteroviruses
View Patent ↗The invention provides cell lines which are useful for the rapid detection of enteroviruses. In particular, the invention provides transgenic African green monkey kidney cell lines and buffalo green monkey kidney cell lines. The invention provides cell lines which have increased sensitivity to infection by enteroviruses in single-cell type and mixed-cell type cultures compared to other cell types which are currently used for enterovirus detection. The cells of the invention also are permissive to infection by a larger number of enteroviruses as compared to the cell type from which they were derived.
1. A transgenic buffalo green monkey kidney cell line expressing human decay accelerating factor (BGMK-hDAF), obtained by a method comprising:
a) transfecting a plurality of buffalo green monkey kidney cells (BGMK) with a vector comprising a sequence encoding human decay accelerating factor (hDAF);
b) contacting said transfected BGMK cells with a selection medium; and
c) selecting transfected BGMK-hDAF cells to obtain a stable transgenic BGMK-hDAF cell line which has a property selected from the group consisting of i) increased sensitivity of detection of one or more enteroviruses compared to a control BGMK cell lacking expression of said hDAF, and ii) permissiveness to echovirus selected from the group consisting of echovirus-6 and echovirus-11; wherein said one or more enteroviruses include polioviruses, Coxsackie A viruses, Coxsackie B viruses, echoviruses, and enterovirus types 68, 69, 70 and 71.
2. The cell line of claim 1 , wherein said human decay accelerating factor is encoded by a sequence selected from SEQ ID NO:1 and SEQ ID NO:3.
3. A composition comprising culture medium and the cell line of claim 1 .
4. The composition of claim 3 , further comprising a cell type other than said transgenic BGMK-hDAF cell line, and wherein said transgenic BGMK-hDAF cell and said cell type are in mixed-cell type culture.
5. The composition of claim 4 , wherein said cell type is selected from the group consisting of RD cells, H292 cells, A549 cells, MRC-5 cells, KB cells and CaCo-2 cells.
6. A method for detection of enterovirus in a sample, comprising:
a) providing i) a sample; and ii) a composition comprising culture medium and a cell of the transgenic BGMK-hDAF cell line of claim 1 ;
b) inoculating said cell with said sample to produce an inoculated cell; and
c) observing said inoculated cell for the presence of said enterovirus.
7. The method of claim 6 , wherein said composition further comprises a cell type other than said transgenic BGMK-hDAF cell line, and wherein said transgenic BGMK-hDAF cell and said cell type are in mixed-cell type culture.
8. The method of claim 7 , wherein said cell type is selected from the group consisting of RD cells, H292 cells, A549 cells, MRC-5 cells, KB cells and CaCo-2 cells.
9. The method of claim 6 , wherein said enterovirus is an echovirus.
10. The method of claim 6 , wherein said enterovirus is one or both of a Coxsackie A virus and a Coxsackie B virus.
11. The method of claim 6 , wherein said enterovirus is a poliovirus.
12. A method for producing enterovirus, comprising:
a) providing i) a sample containing an enterovirus; and ii) a composition comprising culture medium and a cell of a the transgenic BGMK-hDAF cell line of claim 1 ; and
b) inoculating said cell with said sample to produce an inoculated cell, wherein said inoculated cell produces said enterovirus.
13. The method of claim 12 , wherein said composition further comprises a cell type other than said transgenic BGMK-hDAF cell line, and wherein said transgenic BGMK-hDAF cell and said cell type are in mixed-cell type culture.
14. The method of claim 13 , wherein said cell type is selected from the group consisting of RD cells, H292 cells, A549 cells, MRC-5 cells, KB cells and CaCo-2 cells.
15. The method of claim 12 , wherein said enterovirus is an echovirus.
16. The method of claim 12 , wherein said enterovirus is one or both of a Coxsackie A virus and a Coxsackie B virus.
17. The method of claim 12 , wherein said enterovirus is a poliovirus.