Facile method and apparatus for the analysis of biological macromolecules in two dimensions using common and familiar electrophoresis formats
View Patent ↗An ensemble of components and methods are disclosed for utilizing two-dimensional electrophoresis in polyacrylamide or related polymer gels using common, existing and familiar electrophoresis formats and equipment. The disclosed invention makes two-dimensional electrophoresis convenient and easy to use for individuals already using vertical “mini-gel” type systems. The invention discloses the combination of pre-cast disposable gels for both first and second separation dimensions using novel support devices and cassettes that simplify the difficult multiple sample handling and processing steps inherent in ordinary two-dimensional electrophoresis methods. Devices and methods are disclosed in said invention that provide exceptional gel to gel reproducibility.
1. A vertical electrophoresis system comprising:
an electrophoresis chamber,
a buffer core positioned within the electrophoresis chamber; and
a cassette sealed to said buffer core, said cassette comprising a plurality of parallel channels enclosing a plurality of semi-rigid supports and a plurality of pH gradient gels, wherein to one face of each of said semi-rigid supports is covalently attached a pH gradient gel of the plurality of pH gradient gels, wherein each said gel is sealed and isolated from the other gels in a separate laminar space of a channel of the plurality of parallel channels;
wherein each said channel comprises a port at the bottom and top of each laminar space for loading of sample.
2. The cassette of claim 1 , wherein said pH gradient gels are isoelectric focusing gels.
3. The cassette of claim 1 , wherein said cassette is from about 8 to about 10 cm high and about 10 cm wide.
4. The cassette of claim 1 , wherein said pH gradient gels are gel strips.
5. The cassette of claim 1 , wherein said space is from about 4 to about 6 mm wide.
6. The cassette of claim 1 , wherein said pH gradient gels are precast disposable gels.
7. The cassette of claim 1 , wherein said pH gradient gels are tube gels.
8. The cassette of claim 4 , wherein said gel strips are from about 3 to about 5 mm wide.
9. The cassette of claim 4 , wherein said gel strips have a length of from about 80 to about 120 mm.
10. The cassette of claim 4 , wherein said gel strips have a thickness when fully hydrated of from about 0.2 to about 1.0 mm.
11. The cassette of claim 1 , wherein said vertical electrophoresis apparatus is a mini-gel electrophoresis apparatus.
12. A vertical electrophoresis apparatus comprising:
a buffer core; and
a gel strip-manifold assembly sealed to said buffer core, wherein said manifold assembly comprises:
a plurality of semi-rigid supports, wherein to one face of each of said semi-rigid supports is covalently attached each of a plurality of gel strips, wherein the plurality of gel strips are arranged in parallel with a space between them; and
an isolating manifold, comprising a plurality of parallel channels,
wherein each of said gel strips is enclosed by a channel of the plurality of parallel channels and sealed and isolated from the other gel strips in a separate laminar space, wherein each said laminar space is defined by the channel enclosing the strip, wherein each said channel comprises a port at the bottom and top of each laminar space for loading of sample.
13. The gel strip-manifold assembly of claim 12 , wherein said gel strips are isoelectric focusing gels.
14. The assembly of claim 12 , wherein said isolating manifold comprises an elastomeric material.
15. The gel strip-manifold assembly of claim 12 , wherein said space is from about 4 to about 6 mm wide.
16. The gel strip-manifold assembly of claim 12 , wherein said gel strips are precast on said semi-rigid support.
17. The gel strip-manifold assembly of claim 12 , wherein said gel strips are from about 3 to about 5 mm wide.
18. The gel strip-manifold assembly of claim 12 , wherein said gel strips have a length of from about 80 to about 120 mm.
19. The gel strip-manifold assembly of claim 12 , wherein said gel strips have a thickness when fully hydrated of from about 0.2 to about 1.0 mm.
20. The apparatus of claim 12 , wherein said apparatus is a mini-gel electrophoresis apparatus.