Reverse transfection of cell arrays for structural and functional analyses of proteins

• US 20070111201A1
• Filed: 09/08/2005
• Published: 05/17/2007
• Est. Priority Date: 04/30/2001
• Status: Abandoned Application

• Current Assignee: Integral Molecular, Inc.
• Original Assignee: Integral Molecular, Inc.
• Inventors: Doranz, Benjamin
• Application Number:
  US11/222,193
• Publication Number:
  US 20070111201A1
• Time in Patent Office: Days
• US Class Current: 435/5
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12M 23/12 Well or multiwell plates C12M25/04 takes precedence
• CPC Class Codes: C12M 35/02 Electrical or electromagnetic means, e.g. for electroporation or for cell fusion
• CPC Class Codes: C12N 2740/16122 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025

Abstract

The present invention relates to articles and methods for determining the function of genes, gene products, and nucleic acid products. The present invention also relates to identifying ligands and binding partners or proteins and nucleic acid products. The present invention also relates to methods and compositions related to reverse-transfection.

Claims

1. A microwell plate comprising an array of nucleic acid molecules and a transfection reagent wherein said nucleic acid molecules and transfection reagent are frozen in solution.

2. The microwell plate of claim 1, wherein the nucleic acid is a plasmid.

3. The microwell plate of claim 1, wherein the nucleic acid is a virus.

4. The microwell plate of claim 1, wherein said array is a microplate.

5. The microwell plate of claim 1, wherein said array is a slide.

6. The microwell plate of claim 1, wherein said solution comprises a sugar.

7. The microwell plate of claim 1, wherein said solution comprises EDTA.

8. An array comprising an array of nucleic acid molecules and a transfection reagent wherein said array of nucleic acid molecules comprises polymorphisms of a gene and wherein said transfection reagent and nucleic acid molecules are frozen or dried onto a surface.

9. An array comprising an array of nucleic acid molecules and a transfection reagent wherein said nucleic acid molecules encode at least one sensor and, optionally, at least one reporter, wherein said nucleic acid molecules and transfection reagent are frozen or dried on to a surface.

10. The array of claim 9 wherein said sensor comprises an antibody or antibody-like molecule.

11. A method of preparing a microarray for reverse-transfection, comprising an array of nucleic acid molecules comprising:

a) depositing a composition comprising a nucleic acid molecule and a transfection reagent into a well of a microarray; and

b) freezing said microarray.

12. The method of claim 11, where said composition comprising a nucleic acid molecule comprises a transfection reagent.

13. The method of claim 11, where said composition comprises a nucleic acid molecule that can be electroporated into a cell.

14. The method of claim 11, wherein said nucleic acid molecule encodes at least part of a virus.

15. The method of claim 11, wherein said nucleic acid molecule encodes for a protein or a transcription element.

16. The method of claim 15 wherein said nucleic acid molecule comprises a promoter and is operatively linked to a reporter gene.

17. The method of claim 15 wherein said nucleic acid molecule encoding for said protein is mutated.

18. The method of claim 15 wherein said protein is fixed onto a surface.

19. The method of claim 15 wherein said protein is specifically attached to a surface using a tag on the protein.

20. The method of claim 11 wherein each well comprising said nucleic acid molecule comprises a specific mutation in said nucleic acid molecule.

21. The method of claim 11 wherein each well or spot of said microarray comprises a nucleic acid molecule comprising a specific cDNA.

22. A method of introducing a nucleic acid molecule into a cell comprising:

a) thawing a microarray comprising a composition comprising a nucleic acid molecule; and

b) contacting a well of said microarray with said cell under appropriate conditions for entry of the nucleic acid molecule into the cell.

23. A method of mapping a function of a nucleic acid molecule product comprising:

a) contacting a composition comprising an array of nucleic acid molecules with a cell under appropriate conditions for entry of the nucleic acid molecule into the cell; and

b) measuring a function of said nucleic acid molecule product.

24. The method of claim 23 wherein the array comprises different mutations of said nucleic acid product and the function of each product is compared to the wild-type product.

25. A method of preparing an array of viable cells in a multi-well plate comprising

a) thawing a frozen microarray comprising an array of nucleic acid molecules;

and

b) contacting a well in said microarray with said cells.

26. The method of claim 25 wherein said array of nucleic acid molecules encodes a protein or a transcriptional element.

27. The method of claim 26 wherein said array of nucleic acid molecules comprises mutations of said protein or transcription element.

28. The method of claim 25 wherein said array of nucleic acid molecules encodes different cDNAs encoding for different proteins or transcriptional elements.

29. A frozen microarray comprising an array of cells, a nucleic acid molecule, and a transfection reagent.

30. A frozen microarray comprising a nucleic acid molecules, an array of cells, a transfection reagent, or combinations thereof wherein said nucleic acid molecules encodes at least one sensor and, optionally, at least one reporter.

31. A method of selecting a vaccine candidate protein comprising:

a) contacting a composition comprising an array of nucleic acid molecules which encodes variants of said protein with a cell under appropriate conditions for entry of the nucleic acid molecule into the cell and expression of said protein on the cell surface; and

b) measuring the binding of said protein to a first antibody has desirable binding properties and is specific for said protein and, optionally, to a second antibody, wherein said second antibody is has undesirable binding properties, wherein a protein that binds to the first antibody, but not to the second antibody is selected as a vaccine candidate.

32. A method for detecting a pathogen in a sample comprising:

a) contacting said sample with an array of cells comprising ligands for said pathogens, wherein said ligands comprise a signaling mechanism that is activated upon binding of said pathogens to said ligands; and

b) detecting a signal, whereby detection of said signal indicates the presence of said pathogen.

33. A method for detecting a protein in a sample comprising:

a) contacting said sample with an array of cells comprising binding partners for said protein, wherein said binding partners comprise a signaling mechanism that is activated upon binding of said protein to said binding partners; and

b) detecting a signal, whereby detection of said signal indicates the presence of said protein.