Methods and systems for screening using microcapillary arrays
원문보기
IPC분류정보
국가/구분
United States(US) Patent
등록
국제특허분류(IPC7판)
C12N-015/10
G01N-033/68
B01L-003/00
C12N-015/81
G01N-033/53
출원번호
US-0489592
(2017-04-17)
등록번호
US-10227583
(2019-03-12)
발명자
/ 주소
Cochran, Jennifer R.
Chen, Bob
Alford, Spencer Caleb
출원인 / 주소
XCELLA BIOSCIENCES, INC.
대리인 / 주소
Morgan, Lewis & Bockius LLP
인용정보
피인용 횟수 :
0인용 특허 :
58
초록▼
High-throughput methods for screening large populations of variant proteins are provided. The methods utilize large-scale arrays of microcapillaries, where each microcapillary comprises a solution containing a variant protein, an immobilized target molecule, and a reporter element. Immobilized targe
High-throughput methods for screening large populations of variant proteins are provided. The methods utilize large-scale arrays of microcapillaries, where each microcapillary comprises a solution containing a variant protein, an immobilized target molecule, and a reporter element. Immobilized target molecules may include any molecule of interest, including proteins, nucleic acids, carbohydrates, and other biomolecules. The association of a variant protein with a molecular target is assessed by measuring a signal from the reporter element. The contents of microcapillaries identified in the assays as containing variant proteins of interest can be isolated, and cells expressing the variant proteins of interest can be characterized. Also provided are systems for performing the disclosed screening methods.
대표청구항▼
1. A method of screening a population of variant proteins comprising the steps of: providing a microcapillary array comprising a plurality of microcapillaries, each microcapillary comprising a cellular expression system expressing a variant protein, a target molecule immobilized on the surface of a
1. A method of screening a population of variant proteins comprising the steps of: providing a microcapillary array comprising a plurality of microcapillaries, each microcapillary comprising a cellular expression system expressing a variant protein, a target molecule immobilized on the surface of a cell, and a reporter element, wherein the variant protein associates with the immobilized target molecule in the microcapillary with a particular affinity, wherein the cellular expression system is an animal system, a fungal system, a bacterial system, an insect system, or a plant system; andmeasuring a signal from at least one reporter element that indicates association of at least one variant protein with at least one immobilized target molecule to identify at least one microcapillary of interest. 2. The method of claim 1, wherein the variant protein associates with the immobilized target molecule in situ within the microcapillaries. 3. The method of claim 1, wherein the cellular expression system is an animal system. 4. The method of claim 3, wherein the animal system is a mammalian system. 5. The method of claim 1, wherein the cellular expression system is a fungal system. 6. The method of claim 5, wherein the fungal system is a yeast system. 7. The method of claim 1, wherein the cellular expression system is a bacterial system. 8. The method of claim 1, wherein the cellular expression system is an insect system. 9. The method of claim 1, wherein the cellular expression system is a plant system. 10. The method of claim 1, wherein the target molecule is a target protein or polypeptide, a target nucleic acid, a target carbohydrate, or a combination of each. 11. The method of claim 1, wherein the target molecule is a native protein. 12. The method of claim 1, wherein the reporter element is a labeled antibody or other binding molecule, wherein the labeled antibody or other binding molecule localizes to an epitope on the variant protein. 13. The method of claim 12, wherein the labeled antibody or other binding molecule is a fluorescently-labeled antibody or other binding molecule. 14. The method of claim 12, wherein the labeled antibody is a primary or a secondary antibody. 15. The method of claim 1, wherein the reporter element is activated within a cell. 16. The method of claim 15, wherein the reporter element comprises a green fluorescent protein or variant. 17. The method of claim 1, wherein the signal is a fluorescent signal, an absorbance signal, a bright-field signal, or a dark-field signal. 18. The method of claim 1, wherein each microcapillary in the microcapillary array comprises 1 to 5 variant proteins from the population of variant proteins. 19. The method of claim 1, wherein the microcapillary array comprises at least 100,000, at least 300,000, at least 1,000,000, at least 3,000,000, or at least 10,000,000 microcapillaries. 20. The method of claim 1, wherein each microcapillary further comprises an agent to improve viability of the cellular expression system. 21. The method of claim 20, wherein the agent is a growth medium. 22. The method of claim 20, wherein the agent is methylcellulose, dextran pluronic F-68, polyethylene glycol, or polyvinyl alcohol. 23. The method of claim 1, wherein the signal is measured by an optical detector. 24. The method of claim 1, wherein the signal is measured by a microscope. 25. The method of claim 1, further comprising the step of isolating the contents of the microcapillary of interest. 26. The method of claim 25, wherein the contents of the microcapillary of interest are isolated by pulsing the microcapillary of interest with a laser.
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