Screening assays and methods
View Patent ↗Screening assays and methods of performing such assays are provided. In certain examples, the assays and methods may be designed to determine whether or not two or more species can associate with each other. In some examples, the assays and methods may be used to determine if a known antigen binds to an unknown monoclonal antibody.
1. A method of screening a monoclonal antibody comprising:
sealing a moldable slab to a substrate, said moldable slab comprising at least one microwell of less than 100 micrometers in diameter, each of said microwells comprising a single antibody-producing cell that secretes monoclonal antibodies in a volume of 10 nanoliters or less of fluid in said microwell allowing said antibody-producing cell to secrete said monoclonal antibodies in said volume;
exposing said volume of said monoclonal antibodies to at least one defined antigen, and
determining if said monoclonal antibodies binds to said antigen, wherein the steps of the method of screening the monoclonal antibody are performed in less than one day.
2. The method of claim 1 , wherein the method of screening the monoclonal antibody is performed in less than 6 hours.
3. The method of claim 1 , wherein the substrate is pretreated with the defined antigen.
4. The method of claim 1 , wherein the substrate is pretreated with a secondary antibody.
5. The method of claim 1 , wherein the moldable slab comprises a microwell array, wherein at least one microwell of the microwell array comprises the antibody-producing cell that secretes the monoclonal antibody.
6. The method of claim 5 , wherein the microwell array is incubated for less than 24 hours while contacting the substrate.
7. The method of claim 5 , wherein the microwell array is incubated for less than 5 hours while contacting the substrate.
8. The method of claim 5 , wherein the microwell array is incubated for less than 2 hours while contacting the substrate.
9. The method of claim 1 , wherein the monoclonal antibody and the defined antigen associate on the substrate forming a microarray.
10. The method of claim 9 , wherein the microarray on the substrate is removed from the moldable slab and is placed in a reservoir containing a medium.
11. The method of claim 10 , wherein the hydridoma in the microwells is maintained in the medium and the microwell array on the moldable slab contacts a new substrate and forms at least one new microarray.
12. The method of claim 11 , wherein between 5 and 100 new microarrays are formed.
13. The method of claim 1 , wherein the antibody bound to the defined antigen is detected.
14. The method of claim 1 , further comprising determining whether the monoclonal antibody binds to a defined secondary antibody on the substrate.
15. The method of claim 1 , wherein said antigen comprises an intact pathogen.
16. The method of claim 1 , wherein said antigen comprises a cell lysate.
17. The method of claim 15 , wherein said intact pathogen is a bacteria, virus, or fungus.
18. The method of claim 1 , wherein said antibody-producing cell is a hybridoma cell.
19. The method of claim 1 , wherein said antibody-producing cell is a primary B cell.
20. The method of claim 1 , wherein said volume is 1 picoliter to 1 nanoliter.
21. The method of claim 1 , wherein said determining step is carried out by fluorescence detection, mass spectrometry, surface plasmon resonance, or a colorimetric assay.
22. The method of claim 1 , wherein antibody-antigen binding is detected using a fluorescence microscope, fluorimeter, or camera.
23. The method of claim 1 , wherein said antibody-antigen binding is detected using an instrument comprising a charge coupled device, a photomultiplier tube, or a diode array.
24. The method of claim 1 , wherein the steps of the method of screening the monoclonal antibody are performed in less than 12 hours.
25. The method of claim 5 , wherein the monoclonal antibody associates with the substrate to form a microarray.
26. The method of claim 13 , wherein antibody-antigen binding is identified using a detectable marker.
27. The method of claim 26 , wherein the detectable marker comprises a fluorescent label, a colorimetric label, or a radiolabel.
28. The method of claim 26 , wherein the detectable marker is a radiolabel.
29. The method of claim 27 , wherein the radiolabel comprises 3 H, 14 C, 32 P, 33 P, 35 S or 125 I.
30. The method of claim 1 , wherein the cell is retrieved from the moldable slab.