Methods for separation and immuno-detection of biomolecules, and apparatus related thereto

• US 9,696,276 B2
• Filed: 05/17/2013
• Issued: 07/04/2017
• Est. Priority Date: 09/27/2008
• Status: Expired due to Fees

• Current Assignee: Boston Heart Diagnostics Corporation (Eurofins Scientific SE)
• Original Assignee: Boston Heart Diagnostics Corporation (Eurofins Scientific SE)
• Inventors: Asztalos, Bela F., Schaefer, Ernst J.
• Primary Examiner(s): LAM, ANN Y
• Application Number:
  US13/896,765
• Publication Number:
  US 20130248370A1
• Time in Patent Office: 1,509 Days
• Field of Search: None
• CPC Class Codes: C12M 33/00 Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus chemical or physical laboratory apparatus in general B01L, devices for taking cell samples A61B10/0045, withdrawing or distributing predetermined quantities of fluid B01L99/00
• CPC Class Codes: G01N 2333/775 Apolipopeptides
• CPC Class Codes: G01N 2550/00 Electrophoretic profiling, e.g. for proteome analysis
• 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
• CPC Class Codes: G01N 27/44778 on a common gel carrier, i.e. 2D gel electrophoresis
• CPC Class Codes: G01N 2800/32 Cardiovascular disorders
• CPC Class Codes: G01N 2800/50 Determining the risk of developing a disease
• CPC Class Codes: G01N 2800/52 Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/561 Immunoelectrophoresis
• CPC Class Codes: G01N 33/6842 Proteomic analysis of subsets of protein mixtures with reduced complexity, e.g. membrane proteins, phosphoproteins, organelle proteins
• CPC Class Codes: G01N 33/92 involving lipids, e.g. cholesterol , lipoproteins, or their receptors steroid hormones G01N33/743

Abstract

Disclosed are methods and apparatus for separation of biomolecules via two-dimensional gel electrophoresis, methods and apparatus for immunoblotting separated biomolecules, and methods for the use of biomolecules processed via the methods and apparatus of the present invention, including use in a clinical setting. The methods and apparatus for separation of biomolecules via two-dimensional gel comprises vertical agarose gel electrophoresis in the first dimension, and the electrophoresis of a novel non-denaturing 3-35% concave gradient polyacrylamide gel in the second dimension. This novel gel can be cast in a modified gel caster that can facilitate the pouring of multiple gels simultaneously. The methods and apparatus for immunoblotting are useful with any type of immunoblotting, including Western blot, Northern blot, and Southern blot analyzes. These methods and apparatus provide safe, efficient and cost-effective immunoblots, while facilitating the reduction of exposure to toxic or radioactive materials, as well as the disposal of those materials.

Claims

1. A device for immunoblotting, the device comprising:

a plurality of incubation chambers, each comprising a body and configured to contain a plurality of membranes within the body; and

a rotational element for rotating the incubation chamber and defining a rotational axis, wherein the incubation chamber rotates in parallel with the rotational axis.

2. The device of claim 1, wherein the rotational element comprises at least two rolls.

3. The device of claim 2, wherein the at least two rolls are parallel with the incubation chamber.

4. The device of claim 1, wherein said incubation chamber is configured to contain an incubation media.

5. The device of claim 1, wherein the incubation chamber is cylindrical in shape.

6. The device of claim 1, wherein the incubation chamber body comprises at least one opening configured to couple to a cover.

7. The device of claim 1, wherein each of the plurality of incubation chambers are operably associated with the rotational element.

8. The device of claim 1, wherein each of the plurality of incubations chambers are configured to contain a plurality of membranes.

9. The device of claim 1, wherein the plurality of membranes comprises 100 membranes or fewer.

10. The device of claim 1, wherein the incubation chamber is operably coupled to a pumping mechanism.

11. The device of claim 10, wherein the pumping mechanism is a peristaltic pump.

12. The device of claim 11, wherein the peristaltic pump is configured to deliver a fluid into and/or out of said incubation chamber.

13. The device of claim 12, wherein the fluid is selected from the group consisting of incubation media, a washing solution, and a combination thereof.

14. The device of claim 1, wherein the device is configured to perform an immunoblotting technique selected from the group consisting of a Northern blot, Southern blot, and Western blot.

15. An assembly for immunoblotting, the assembly comprising

a plurality of incubation chambers, each incubation chamber comprising a body and configured to contain a plurality of membranes within the body;

and

a rotational element for rotating the plurality of incubation chambers and defining a rotational axis, wherein each incubation chamber rotates in parallel with the rotational axis.

16. The assembly of claim 15, wherein the rotational element comprises at least two rolls.

17. The assembly of claim 16, wherein the at least two rolls are parallel with the plurality of incubation chambers.

18. The assembly of claim 15, wherein the plurality of incubation chambers is operably coupled to a pumping mechanism.

19. The assembly of claim 18, wherein the pumping mechanism comprises a peristaltic pump configured to deliver a fluid into and/or out of the incubation chamber.