Composite compositions for electrophoresis

• US 20050121325A1
• Filed: 09/20/2004
• Published: 06/09/2005
• Est. Priority Date: 09/19/2003
• Status: Abandoned Application

• Current Assignee: Life Technologies Israel Ltd. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Life Technologies Israel Ltd. (Thermo Fisher Scientific Incorporated)
• Inventors: Margalit, Ilana, Thacker, Michael, Updyke, Timothy
• Application Number:
  US10/946,472
• Publication Number:
  US 20050121325A1
• Time in Patent Office: Days
• US Class Current: 204/469
• CPC Class Codes: G01N 27/44739 Collecting the separated zones, e.g. blotting to a membrane or punching of gel spots
• CPC Class Codes: G01N 27/44747 Composition of gel or of carrier mixture

Abstract

The invention is drawn to composite agarose/acrylamide compositions and gels. In particular it relates to gels for the separation of molecules, particularly macromolecules such as proteins. The invention is also directed to the preparation of composite gels, the separation of molecules by techniques such as electrophoresis using such gels, and the transfer of proteins from such gels to a transfer membrane using an immunoblot transfer gel.

Claims

1. A composition comprising agarose, polyacrylamide and a photoinitiator.

2. The composition of claim 1, further comprising one or more components selected from the group consisting of one or more salts, one or more ions, and one or more denaturants.

3. The composition of claim 1, wherein said composition is in a gel format.

4. The composition of claim 3, wherein said gel format is selected from a group consisting of a 96-well gel format and a 48-well gel format.

5. The composition of claim 3, wherein said gel format is a 96-well staggered gel format.

6. A gel comprising agarose, polyacrylamide and a photoinitiator.

7. The gel of claim 6, wherein said gel is an electrophoretic separation gel.

8. The separation gel of claim 7, wherein said gel is a pre-cast gel.

9. The separation gel of claim 7, wherein said gel is an E-PAGE™

96 Gel substantially as described herein.

10. A method of resolving macromolecules, comprising subjecting said macromolecules to electrophoresis through the separation gel of claim 7.

11. The method of claim 10, wherein said macromolecules are proteins.

12. A kit comprising the separation gel of claim 6 or the pre-cast separation gel of claim 8.

13. The kit of claim 12, further comprising one or more sample loading buffers.

14. The kit of claim 12, further comprising one or more sample buffers for electrophoresis.

15. The kit of claim 12, further comprising one or more protein standards.

16. The kit of claim 12, further comprising one or more sets of instructions.

17. The kit of claim 12, further comprising one or more pre-cut membranes for use in blotting, said pre-cut membranes having a length and a width, wherein the pre-cut membranes are pre-cut to match the length and width as the pre-cast gel.

18. The kit of claim 12, further comprising an immunoblot transfer gel.

19. The kit of claim 18, wherein the transfer gel comprises agarose.

20. The kit of claim 18, wherein the gel comprises between 0.5% and 2% agarose.

21. The kit of claim 18, wherein the separation gel comprises surface irregularities on a top gel surface.

22. The kit of claim 18, wherein the immunoblot transfer gel comprises immunoblot transfer buffer.

23. A system for electrophoresis, said system comprising:

(a) a gel comprising agarose, polyacrylamide and a photoinitiator; and

(b) a power supply comprising a power regulator.

24. The system for electrophoresis of claim 23, wherein said power regulator provides constant power over a period of time sufficient for a set of proteins to resolve.

25. The system for electrophoresis of claim 24, wherein said set of proteins comprises at least two proteins having molecular weights selected from the group consisting of 20 kDa, 40 kDa, 60 kDa, 120 kDa and 220 kDa.

26. The system for electrophoresis of claim 23, wherein said power supply is an E-Base™

power supply substantially as described herein.

27. The system for electrophoresis of claim 23, wherein said system is a high throughput system.

28. A gel comprising:

agarose;

polyacrylamide;

a photoinitiator; and

BES.

29. The gel of claim 28, wherein the agarose is present at a concentration of between 1% and 2% and the BES is present at a concentration of between 10 mM and 250 mM.

30. The gel of claim 28, wherein the gel has a first layer, a second layer, and a third layer, each layer comprising:

agarose; and

polyacrylamide, wherein said second layer further comprises a photoinitiator.

31. The gel of claim 28, further comprising an amine transfer agent.

32. An immunoblot transfer gel comprising agarose and immunoblot transfer buffer.

33. The immunoblot transfer gel of claim 32, comprising 0.5% to 2% agarose.

34. The immunoblot transfer gel of claim 32, wherein the immunoblot transfer buffer comprises Tris buffer and methanol.