In one aspect the present invention provides a method of identifying a cell or cell colony which produces a polypeptide of interest, the method comprising exposing one or more cells to a marker compound which associates with a reference polypeptide, wherein production of the polypeptide of interest
In one aspect the present invention provides a method of identifying a cell or cell colony which produces a polypeptide of interest, the method comprising exposing one or more cells to a marker compound which associates with a reference polypeptide, wherein production of the polypeptide of interest by the cells is linked to production of the reference polypeptide, and detecting association of the marker compound with the one or more cells, thereby identifying a cell or cell colony which produces the polypeptide of interest.
대표청구항▼
1. A method of identifying a cell or cell colony which produces a polypeptide of interest, the method comprising: a) exposing one or more cells in a same medium to tetrahydrofolate and a marker compound comprising methotrexate which associates with a reference polypeptide comprising dihydrofolate re
1. A method of identifying a cell or cell colony which produces a polypeptide of interest, the method comprising: a) exposing one or more cells in a same medium to tetrahydrofolate and a marker compound comprising methotrexate which associates with a reference polypeptide comprising dihydrofolate reductase, wherein production of the polypeptide of interest by the one or more cells is linked to production of the reference polypeptide; andb) detecting association of the marker compound with the one or more cells, thereby identifying a cell or cell colony which produces the polypeptide of interest,wherein a gene encoding the reference polypeptide is amplified in the identified cell or cell colony, andwherein detecting association of the marker compound comprises (i) obtaining an image of the one or more cells and (ii) analysing the image to identify association of the marker compound with the identified cell or cell colony. 2. A method according to claim 1, wherein a gene encoding the polypeptide of interest is amplified in the identified cell or cell colony. 3. A method according to claim 1, wherein the polypeptide of interest comprises a biotherapeutic molecule. 4. A method according to claim 1, wherein the polypeptide of interest comprises an immunoglobulin or fragment thereof. 5. A method according to claim 1, wherein the polypeptide of interest comprises a receptor. 6. A method according to claim 1, wherein the polypeptide of interest comprises a recombinant polypeptide expressed by a transfected cell. 7. A method according to claim 1, wherein the polypeptide of interest and the reference polypeptide are encoded by one or more exogenous vectors introduced into the one or more cells. 8. A method according to claim 1, wherein the one or more cells are cultured cells. 9. A method according to claim 1, wherein the image is obtained and analysed by an automated imaging system. 10. A method according to claim 1, wherein the image is analysed to determine a level of the marker compound associated with the cell or cell colony, the level being indicative of an amount of the polypeptide of interest produced by the cell or cell colony. 11. A method according to claim 1, further comprising a step of exposing the one or more cells to an agent which binds to the polypeptide of interest and detecting binding of the agent to the one or more cells. 12. A method according to claim 1, wherein the one or more cells are pre-incubated with a compound comprising methotrexate. 13. A method according to claim 1, wherein the one or more cells are disposed on or in solid or semi-solid medium. 14. A method according to claim 1, wherein the polypeptide of interest is secreted by the one or more cells. 15. A method according to claim 1, wherein the reference polypeptide is an intracellular or cell surface-associated polypeptide. 16. A method according to claim 1, wherein the marker compound comprises a fluorescent label. 17. A method according to claim 16, wherein the fluorescent label comprises fluorescein, rhodamine or phycoerythrin. 18. A method of selecting a cell or cell colony which produces a polypeptide of interest from a plurality of cells, comprising detecting a cell or cell colony which produces the polypeptide of interest by a method according to any preceding claim, and selecting the cell or cell colony by picking the detected cell or cell colony. 19. A method according to claim 18, comprising: a) determining a level of the marker compound associated with each cell or cell colony;b) comparing the level to a predetermined threshold; andc) selecting a cell or cell colony having a level of the marker compound above the predetermined threshold. 20. A method according to claim 18, wherein a cell or cell colony which is associated with an elevated level of the marker compound is selected, thereby selecting a cell or cell colony showing an elevated amount of production of the polypeptide of interest. 21. A method according to claim 20, wherein the level of the marker compound in the selected cell or cell colony is elevated relative to a mean level of the marker compound in the one or more cells. 22. A method according to claim 18, wherein the cell or cell colony is picked by an automated cell picking device. 23. A method according to claim 1, wherein exposing the one or more cells to a marker compound is performed at least in part with the marker compound disposed in a semi-solid medium and with the one or more cells disposed in or on the semi-solid medium, and wherein obtaining an image of the one or more cells is performed while the one or more cells are still disposed in or on the semi-solid medium. 24. A method according to claim 1, wherein detecting association of the marker compound further comprises picking the cell or cell colony from a plurality of cells, and wherein the cell or cell colony picked exhibits an elevated level of association of the marker compound relative to a mean level of the marker compound in the one or more cells. 25. A method according to claim 1, wherein the one or more cells are exposed to an amount of tetrahydrofolate effective to mitigate a toxicity of the marker compound. 26. A method of identifying a cell or cell colony which produces an amplifiable marker polypeptide comprising dihydrofolate reductase, the method comprising: (a) exposing one or more cells in a same medium to tetrahydrofolate and a marker compound comprising methotrexate which associates with the amplifiable marker polypeptide, the amplifiable marker polypeptide comprising a product of a gene which can be amplified in the one or more cells in the presence of the marker compound; and(b) detecting association of the marker compound with the one or more cells, thereby identifying a cell or cell colony which produces the amplifiable marker polypeptide,wherein detecting association of the marker compound comprises (i) obtaining an image of the one or more cells and (ii) analyzing the image to identify association of the marker compound with the identified cell or cell colony.
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이 특허에 인용된 특허 (1)
David Gary S. (La Jolla CA) Greene Howard E. (Carlsbad CA), Immunometric assays using monoclonal antibodies.
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