Automated system for two-dimensional electrophoresis
First Claim
Patent Images
1. An apparatus for preparing a gel medium for separating a plurality of protein molecules, said apparatus comprising:
- a) a movable mold half which, when placed near a gel-binding material, forms a mold cavity;
b) means for moving said movable mold half;
c) means for transporting said gel-binding material; and
d) means for dispensing a polymerizable gel mixture into said mold cavity,wherein said means for dispensing said polymerizable gel mixture comprises a precision gradient maker, wherein said gradient maker comprises a plurality of syringes coupled to a plurality of valves and said valves are coupled to a plurality of reservoirs.
1 Assignment
0 Petitions
Accused Products
Abstract
The present invention provides an integrated, fully automated, high-throughput system for two-dimensional electrophoresis comprised of gel-making machines, gel processing machines, gel compositions and geometries, gel handling systems, sample preparation systems, software and methods. The system is capable of continuous operation at high-throughput to allow construction of large quantitative data sets.
176 Citations
74 Claims
-
1. An apparatus for preparing a gel medium for separating a plurality of protein molecules, said apparatus comprising:
-
a) a movable mold half which, when placed near a gel-binding material, forms a mold cavity; b) means for moving said movable mold half; c) means for transporting said gel-binding material; and d) means for dispensing a polymerizable gel mixture into said mold cavity, wherein said means for dispensing said polymerizable gel mixture comprises a precision gradient maker, wherein said gradient maker comprises a plurality of syringes coupled to a plurality of valves and said valves are coupled to a plurality of reservoirs. - View Dependent Claims (2, 3, 4, 5, 6, 7)
-
-
8. A method of preparing a gel medium for separating a plurality of protein molecules, wherein said method comprises steps of:
-
a) attaching a gel-binding material to a longitudinal mold cavity; b) dispensing a variable composition of a polymerizable gel mixture into said mold cavity through a delivery tube having an open end, wherein said open end is inserted into said mold cavity, wherein said gel mixture comprises a gradient and wherein said gel mixture becomes chemically bonded to said backing material; c) withdrawing said delivery tube from said mold cavity during said dispensing whereby said open end of said delivery tube remains at or just above the rising meniscus of said gel mixture in said mold cavity; and d) removing said gel-binding material and said gel mixture from said mold cavity, wherein said gradient is dispensed as part of a sequence of segments comprising; i) a first segment similar in volume to the volume of said delivery tube, wherein said first segment is dispensed before insertion of said delivery tube into said mold cavity; ii) a second segment comprising the remainder of said gradient, wherein said second segment is dispensed after insertion of said delivery tube into said mold; and iii) a third segment having a volume similar to the volume of said first segment.
-
-
9. A method of separating a plurality of proteins wherein said proteins are in a liquid sample, said method comprising steps of:
-
a) applying said liquid sample onto a surface of a gel medium, wherein said gel medium comprises a pH gradient and is attached to a backing material; b) applying a voltage across said gel medium to effect a separation of said proteins; and c) subjecting said gel medium to conditions wherein the volume of said liquid sample is substantially decreased through inhibition of said liquid sample into said gel medium or through loss of water in said liquid sample to an insulating gaseous or a liquid environment of said gel, such that said proteins are substantially incorporated into said gel. - View Dependent Claims (10)
-
-
11. An apparatus for processing a gel medium for separating a plurality of protein molecules wherein said gel medium is fixed to a backing material, wherein said apparatus comprises:
-
a) a plurality of stations; b) means for loading said gel medium with said protein molecules; c) means for reversibly grasping said backing material; and d) means for transporting said gel medium to said stations, wherein one or more of said stations comprise means for application of a voltage longitudinally across said gel medium. - View Dependent Claims (12)
-
-
13. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises:
-
a) a first segment, wherein said first segment is planar and provides a medium for separating said protein molecules; and b) a second segment, wherein said second segment is of greater thickness than said first segment and comprises a buffer reservoir for supplying ions, wherein a rigid electrode is embedded within said second segment and wherein said electrode is used to apply a voltage across said gel. - View Dependent Claims (14)
-
-
15. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises:
-
a) a first segment, wherein said first segment is planar and provides a medium for separating said protein molecules; and b) a second segment, wherein said second segment is of greater thickness than said first segment and comprises a buffer reservoir for supplying ions, wherein an internal slot is formed in said second segment during molding, the floor of said slot being at or near a junction between said first segment and said second segment.
-
-
16. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises:
-
a) a first segments wherein said first segment is planar and provides a medium for separating said protein molecules; and b) a second segment wherein said second segment is of greater thickness than said first segment and comprises a buffer reservoir for supplying ions, and c) a third segment, wherein said third segment is interposed between said first segment and said second segment, said third segment is a stacking gel, wherein said protein molecules are stacked between a first low molecular weight ionic species and a second low molecular weight ionic species prior to effecting a separation of said protein molecules in said first segment.
-
-
17. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises:
-
a) a first segment, wherein said first segment is planar and provides a medium for separating said protein molecules; and b) a second segment, wherein said second segment is of greater thickness than said first segment and comprises a buffer reservoir for supplying ions, and c) a third segment, wherein said third segment is interposed between said first segment and said second segment, and wherein said third segment comprises a composition different from the compositions of said first segment and said second segment and wherein said third segment has greater strength and elasticity than said first segment and said second segment.
-
-
18. A method of performing electrophoresis in an acrylamide gel, said method comprises steps of:
-
a) polymerizing said gel in a mold; b) removing said gel from said mold by mechanical interaction with a region of said gel; and c) performing an electrophoretic separation in said gel. - View Dependent Claims (19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 72)
-
- 46. A method of detecting macromolecules in an electrophoresis gel which method comprises the step of suspending said gel by an edge of said gel in a solution or a sequence of solutions such that said macromolecules are rendered detectable.
-
54. A method for scanning a stained gel medium, wherein said method comprises the steps of:
-
a) grasping an edge of said gel medium; b) transporting said gel medium to a scanning station by a mechanical motion means; and c) placing said gel medium in a space illuminated by a light source and within view of a light detector. - View Dependent Claims (55, 56, 57, 58, 59)
-
-
60. A method for separating a plurality of protein molecules in a plurality of gel mediums, said method comprising steps of:
-
a) grasping an edge of each of said gel mediums; b) transporting each of said gel mediums to a plurality of stations wherein said stations comprise a plurality of environments; and c) inserting each of said gel mediums into said stations for separating said protein molecules. - View Dependent Claims (61, 62)
-
-
63. A programmable system for separating a plurality of protein molecules in a plurality of gel mediums respectively, said system comprising:
-
a) means for preparing said gel mediums; b) means for loading each of said gel mediums with said protein molecules; c) means for separating said protein molecules in each of said gel mediums; d) means for staining or otherwise revealing proteins in each of said gels; and e) means for scanning each of said gel mediums to detect resolved macromolecules, wherein said programmable system comprises a plurality of stations, further wherein each of said gel mediums is assigned thereto a plurality of parameters for separating said protein molecules in each of said gel mediums, and further comprising means for grasping each of said gel mediums at each of said stations.
-
-
64. A programmable system for separating a plurality of protein molecules in a plurality of gel mediums respectively, said system comprising:
-
a) means for preparing said gel mediums; b) means for loading each of said gel mediums with said protein molecules; c) means for separating said protein molecules in each of said gel mediums; d) means for staining or otherwise revealing proteins in each of said gels; and e) means for scanning each of said gel mediums to detect resolved macromolecules. wherein said programmable system comprises a plurality of stations, further wherein each of said gel mediums is assigned thereto a plurality of parameters for separating said protein molecules in each of said gel mediums, further comprising means for transporting each of said gel mediums to said stations.
-
-
65. An integrated system for two-dimensional electrophoresis, said system comprising:
-
a) means for preparing an isoelectric focusing gel; b) means for loading said isoelectric focusing gel with a plurality of protein molecules; c) means for applying a first voltage across said isoelectric focusing gel; d) means for preparing a slab electrophoresis gel; e) means for loading said isoelectric focusing gel onto said slab electrophoresis gel; f) means for applying a second voltage across said slab electrophoresis gel; and g) means for staining said slab electrophoresis gel. - View Dependent Claims (66, 67, 68, 69, 70)
-
-
71. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises two regions of distinct geometry wherein a first region is essentially planar and provides a medium for separating a plurality of macromolecules and a second region is nonplanar and provides a means for suspending said gel by an edge in liquid or gaseous surroundings, and wherein part of said gel medium is polymerized around a rigid support.
-
73. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises three regions of distinct geometry:
-
a first region that is essentially planar and provides a medium for separating a plurality of macromolecules, a second region is nonplanar and provides a means for suspending said gel by an edge in liquid or gaseous surroundings, and a third region that is interposed between said first and second region, and wherein said third region has a different composition than said first and second region and said third region displays greater strength and elasticity than said first and second region.
-
-
74. A gel medium for separating a plurality of protein molecules, wherein said gel medium comprises two regions of distinct geometry wherein a first region is essentially planar and provides a medium for separating a plurality of macromolecules and a second region is nonplanar and provides a means for suspending said gel by an edge in liquid or gaseous surroundings, and wherein a rigid electrode is polymerized within said second region, said electrode extends outside of said second region at one or more locations and said electrode is used to apply a voltage across said gel medium.
Specification