Continuous gel casting method and apparatus

• US 6,187,250 B1
• Filed: 08/19/1998
• Issued: 02/13/2001
• Est. Priority Date: 08/19/1998
• Status: Expired due to Fees

• Current Assignee: James Champagne
• Original Assignee: James Champagne
• Inventors: Champagne, James T.
• Primary Examiner(s): Vargot, Mathieu D.
• Application Number:
  US09/136,525
• Time in Patent Office: 909 Days
• Field of Search: 264/299, 264/102, 264/494, 264/495, 264/496, 264/40.6, 264/145, 264/148, 425/174.4, 425/371, 425/364.R, 425/143, 425/144, 425/145, 425/308, 425/131.1, 425/812, 204/467, 204/469, 204/470, 204/619, 204/620, 204/621
• US Class Current: 264/495
• CPC Class Codes: B29C 39/16 between endless belts
• CPC Class Codes: B29K 2105/0061 Gel or sol
• CPC Class Codes: G01N 27/44704 Details; Accessories
• CPC Class Codes: Y10S 425/812 Venting

Abstract

An apparatus and method which allows one skilled in the art to make either gradient or non-gradient slab gels continuously so that the produced gels are uniformly formed, polymerized and cut to a specific size as needed, in a mass-produced, assembly line manner.

Claims

1. An apparatus for the continuous casting of gels comprising:

a) a means of introducing into a molding space a reaction mixture capable of becoming a gel, wherein said means of introducing into the molding space the reaction mixture, comprises at least one incremental formulation reservoir that delivers pre-formulated reaction mixture solution through an equal number of nozzles into one vertical edge of said molding space in order to create predetermined gradient conditions, and said reaction mixture solution is delivered to a sealed molding space from at least one incremental formulation reservoir at one vertical edge of the molding space;

b) a casting manifold enclosing said molding space and said casting manifold providing a means of creating a continuously moving, sealed molding space for maintaining said reaction mixture during a gelation process;

c) a means for formulating a vertical gradient of composition in a plurality of components within said reaction mixture as said reaction mixture enters said molding space by pre-formulating a composition of an incremental reaction mix formula;

d) a venting means to displace air entrained in the reaction mixture within the molding space;

e) a means of initiation of a polymerization process as said reaction mixture enters said molding space;

f) a means of controlling the temperature of the reaction mixture during the gelation process as the reaction mixture travels through the apparatus;

g) a means for removing a polymerized gel from the molding space;

h) a means for cutting the gel to a desired length;

i) a means for removing excess gel formed during the process; and

j) a means for stacking the polymerized, cut gels.

2. The apparatus of claim 1, wherein, said means of introducing into the molding space the reaction mixture comprises a multi-chamber cylinder block and pistons, which allows said reaction mixture solutions to be delivered in a controlled and uniform rate.

3. The apparatus of claim 2, wherein said multi-chamber cylinder block and pistons further comprises:

a) a series of layered metal, plastic or other composite laminations having an array of aligned holes that when laminated together vertically, define a plurality of vertical cylinders or cylindrical spaces when stacked together;

b) said plurality of cylinders having an inlet end and an outlet end, and each cylinder having a tight fitting glass or plastic column sleeve inserted with O-ring seals at the inlet and outlet end of said column, and which terminate at the end of the cylinder and close the cylinder against a specific inlet and outlet layer of said multi-chamber cylinder block so that it defines a primary reservoir, said primary reservoirs terminating in individual pairs of one inlet and one outlet layer of said multi-chamber cylinder block, and each primary reservoir is provided at least one nozzle at the molding space and a horizontal channel in said outlet layer that defines an outlet channel, and at least one piston is disposed within each primary reservoir and separates the primary reservoirs into an inlet and an outlet chamber such that said reaction mix formula resides within the outlet chamber connected to the outlet channel, and a horizontal inlet channel is provided in said inlet layer of each reservoir so that they connectively attach each primary reservoir to a common inlet channel in order to provide a positive hydraulic pressure, and within said common inlet chamber is a drive fluid and said inlet chamber is connected to a common pump that supplies a uniform pressure to all of the primary reservoirs and a uniform flow rate for each primary reservoir;

c) the primary reservoirs in said multi-chamber cylinder block are arranged such that those incremental formulation reservoirs that supply reaction mix formula to a topmost one-half of said molding space are arranged so that their inlet ends are above said molding space and those incremental formulation reservoirs that supply reaction mix formula to a lowermost one-half of said molding space are arranged so that their inlet ends are below said molding space; and

d) a means of calibrating and adjusting a plurality of relative flow rates between primary reservoirs in order to keep all incremental reaction mix formulation flow rates and overall volumes equivalent over the entire production run.

4. The apparatus of claim 3, wherein said means of calibrating and adjusting the relative flow rates between primary reservoirs comprises:

a) an orifice plate positioned at the horizontal outlet layer of each primary reservoir and disposed between said outlet chamber of said primary reservoir and said horizontal outlet layer;

b) said orifice plate having at least three distinct laminated layers such that an outer two layers of said orifice plate are comprised of rigid materials selected from metal, or plastic or other laminate, and a middle layer of said orifice plate is comprised of a compressible elastomeric material;

c) a plurality of transversing channels penetrating at least two layers of said orifice plate and where one of the penetrated layers being said middle layer, thereby connecting said outlet chamber of said primary reservoir to said horizontal outlet channel;

d) a screw-cap means positioned at the horizontal inlet layer for each primary reservoir and disposed between an inlet chamber of said primary reservoir and said horizontal inlet channel, and connectively extending through said horizontal inlet layer to a surface of said multi-chamber cylinder block such that said screw-cap can be threadedly affixed to said cylinder block through the inlet layer of said cylinder block;

e) said screw-cap means also having a plurality of transverse channels connecting said inlet chamber of said primary reservoir to said horizontal inlet channel such that said plurality of transverse channels remain connected between said inlet chamber of said primary reservoir and said horizontal inlet channel as said screw-cap means is adjusted and vertically repositioned relative to said multi-chamber cylinder block;

f) a means to seal said screw-cap means between said transverse channels in said inlet layer and said multi-chamber cylinder block; and

g) said screw-cap means being capable of generating a compressive force transmitted to said orifice plate and can compress said compressible elastomeric middle layer of said orifice plate.

5. The apparatus of claim 4 wherein said casting manifold comprises:

a) a cavity having two sides comprised of a plurality of flexible moving belts, a top and bottom, and an upstream and a downstream end;

b) wherein said top and bottom of said cavity are each sealed by an opposable compression roller means comprised of at least two opposed compression rollers, a lower and an upper, along a length of the top and bottom of said molding space, and having a suitable vertical width so as to form a seal under less than 1 atmosphere pressure, wherein said lower opposable compression roller supports the gel introduced into said molding space onto a secondary support means such that the secondary support means is also supported within said molding space by the opposable compression roller means;

c) said plurality of flexible moving belts having a continuous mating surface on the cavity forming face of said flexible moving belts at the top and bottom of said molding space where said opposable compression roller means seal said molding space, and are capable of being compressed closed by said opposable compression roller means to create the seal as the flexible moving belts enter a gap between a pair of rigid platens, said rigid platens are each positioned adjacent to said flexible moving belts and on the opposite side of each belt from the side which is in contact with the reaction mixture on each side of the molding space;

d) the upstream end of the casting cavity being defined by the end wherein the reaction solution is introduced into said molding space, and the downstream end of this cavity being defined by the end of the cavity where a solidifying gel hardens as it moves through the molding space, and wherein the length of the casting cavity is sufficient to provide time enough for the reaction mixture to solidify within in the molding space at a particular speed that said flexible moving belts are moving; and

e) said flexible moving belts being fitted around a series of drive roller means and idling roller means which can also act as compression roller means that seal said molding space, that maintain said mated belts at a constant speed as they move downstream during a polymerization process.

6. The apparatus of claim 5, wherein said secondary support means comprises a backing selected from the group consisting of:

a rigid laminate, a mask, glass, or plastic.

7. The apparatus of claim 6, wherein both rigid platens have a finely adjustable gap between them, wherein said gap does not vary in width more than an acceptable amount so that a thickness of the solidifying reaction mixture is maintained during the gelation process by opposing internal manifold pressure that opposably applies pressure to the belts against said rigid platens.

8. The apparatus of claim 7 wherein said mating surfaces of the upper and lower belts have a bead on one belt and a groove to sealingly receive the bead on the other belt in a “

ziplock”

style configuration.

9. The apparatus of claim 8 wherein said top and bottom of said cavity are sealed by an upper and lower belt means wherein said lower belt means supports the gel introduced into said molding space onto the secondary support means and sealing along the lateral edges of the upper and lower belts is obtained through application of pressure onto the flexible and resilient belts.

10. The apparatus of claim 9, wherein the thickness of the solidifying reaction mixture is in the range from about 0.15 mm to about 5 mm.

11. The apparatus of claim 9, wherein the thickness of the solidifying gel is about 1 mm.

12. The apparatus of claim 9, wherein said rigid platens also provide temperature control during the gelation process.

13. The apparatus of claim 12, wherein said rigid platens also provide a means for illuminating the reaction mixture with UV light during the gelation process.

14. The apparatus of claim 12, wherein said venting means further comprises a means of collecting any entrained air displaced by the reaction mixture as it flows out of holes or other cavities, in a secondary support or a mask, and within said molding space where said secondary support enters at the upstream end of said molding space, and on a side of said secondary support away from a side where the reaction mixture is introduced.

15. The apparatus of claim 14, wherein said venting means further comprises a small chamber filled with reaction mixture at a pressure that is just slightly below a pressure in the main cavity, where the reaction mixture is introduced, so that any displaceable entrained air bubbles in the reaction mixture in the molding space can be removed as they float up and collect together into an air bell means positioned adjacent to said molding space and said secondary support at a top of a cavity, said air bell means having a release valve that can vent the air as it builds up while maintaining a constant pressure within the molding space.

16. The apparatus of claim 15, wherein said venting means has a level detector means located adjacent to a small chamber, and said level detector means maintains the air/liquid interface at a predetermined level by releasing air out the top of the collector chamber through a release valve.

17. The apparatus of claim 16, wherein said level detector means is designed with an adjustable “

dead-band”

or hysteresis in the level of response to filter out short term variation in flow rate.

18. The apparatus of claim 16, wherein said wherein said means of initiation of the polymerization process is through a means of chemical generation of free radicals, wherein said chemical generation means further comprises:

a) a series of layered metal, plastic or other composite laminations having an array of aligned holes that when laminated together vertically, define a plurality of vertical cylinders or cylindrical spaces when stacked together;

b) said plurality of cylinders having an inlet and an outlet end, and each cylinder having a tight fitting glass or plastic column sleeve inserted with O-ring seals at the inlet and outlet end of said column, and which terminate at the end of the cylinder and close the cylinder against a specific inlet and outlet layer of said multi-chamber cylinder block so that it defines a primary reservoir, said primary reservoirs terminating in individual pairs of one inlet and one outlet layer of said multi-chamber cylinder block, and each primary reservoir is provided at least one nozzle at the molding space and a horizontal channel in said outlet layer that defines an outlet channel, and at least one piston is disposed within each primary reservoir and separates the primary reservoirs into an inlet and an outlet chamber such that said reaction mix formula resides within the outlet chamber connected to the outlet channel, and a horizontal inlet channel is provided in said inlet layer of each reservoir so that they connectively attach each primary reservoir to a common inlet channel in order to provide a positive hydraulic pressure, and within said common inlet chamber is a drive fluid and said inlet chamber is connected to a common pump that supplies a uniform pressure to all of the primary reservoirs and a uniform flow rate for each primary reservoir;

c) the primary reservoirs in said multi-chamber cylinder block are arranged such that those incremental formulation reservoirs that supply reaction mix formula to a topmost one-half of said molding space are arranged so that their inlet ends are above said molding space and those incremental formulation reservoirs that supply reaction mix formula to a lowermost one-half of said molding space are arranged so that their inlet ends are below said molding space; and

d) a means of calibrating and adjusting a plurality of relative flow rates between primary reservoirs in order to keep all incremental reaction mix formulation flow rates and overall volumes equivalent over the entire production run.

19. The apparatus of claim 18, wherein said means of calibrating and adjusting the relative flow rates between primary reservoirs comprises:

a) an orifice plate positioned at the horizontal outlet layer of each primary reservoir and disposed between said outlet chamber of said primary reservoir and said horizontal outlet layer;

b) said orifice plate having at least three distinct laminated layers such that an outer two layers of said orifice plate are comprised of rigid materials selected from metal, or plastic or other laminate, and a middle layer of said orifice plate is comprised of a compressible elastomeric material;

c) a plurality of transversing channels penetrating at least two layers of said orifice plate and where one of the penetrated layers being said middle layer, thereby connecting said outlet chamber of said primary reservoir to said horizontal outlet channel;

d) a screw-cap means positioned at the horizontal inlet layer for each primary reservoir and disposed between an inlet chamber of said primary reservoir and said horizontal inlet channel, and connectively extending through said horizontal inlet layer to a surface of said multi-chamber cylinder block such that said screw-cap can be threadedly affixed to said cylinder block through the inlet layer of said cylinder block;

e) said screw-cap means also having a plurality of transverse channels connecting said inlet chamber of said primary reservoir to said horizontal inlet channel such that said plurality of transverse channels remain connected between said inlet chamber of said primary reservoir and said horizontal inlet channel as said screw-cap means is adjusted and vertically repositioned relative to said multi-chamber cylinder block;

f) a means to seal said screw-cap means between said transverse channels in said inlet layer and said multi-chamber cylinder block; and

g) said screw-cap means being capable of generating a compressive force transmitted to said orifice plate and can compress said compressible elastomeric middle layer of said orifice plate.

20. The apparatus of claim 19, wherein said chemical generation means further comprises:

a) a series of layered plastic or other composite laminations having an array of aligned holes that when laminated together vertically, define vertical a plurality of cylindrical spaces or cylinders when stacked together;

b) said plurality of cylinders having a inlet and outlet end, and each cylinder having a tight fitting glass or plastic column sleeve inserted with O-ring seals at an inlet and an outlet end of said column, and which terminate at the end of the cylinder and close the cylinder against a specific inlet and outlet layer of a multi-chamber cylinder block so that it defines a primary reservoir, said primary reservoirs terminating in individual pairs of one inlet and one outlet layer of said multi-chamber cylinder block, and each primary reservoir is provided at least one nozzle at the molding space and a horizontal channel in said outlet layer that defines an outlet channel, and at least one piston is disposed within each primary reservoir and separates the primary reservoirs into an inlet and an outlet chamber such that said reaction mix formula resides within the outlet chamber connected to the outlet channel, and a horizontal inlet channel is provided in said inlet layer of each reservoir so that they connectively attach each primary reservoir to a common inlet channel in order to provide a positive hydraulic pressure, and within said inlet chamber is a drive fluid and said inlet chamber is connected to a common pump that supplies a uniform pressure to all of the primary reservoirs and a uniform flow rate for each primary reservoir;

c) a secondary reservoir having a diameter that is less than about ⅔

of a diameter of the primary reservoir;

d) said secondary reservoir positioned concentrically within the primary reservoir so that the primary glass column and a secondary glass column both have top ends terminating in adjacent multi-layer cylinder block layers and both having outlet channels terminating in the same cylinder block layer;

e) said primary and secondary glass columns both having spacer rings disposed between said primary and secondary glass columns and between the primary glass column and the cylinder block;

f) said cylinder block layers are separated by a gasket means having disposed within said gasket means, horizontal channels connecting said primary and secondary reservoir contents to a nozzle;

g) said cylinder block layers also having a circular opening in which a removable concentric orifice ring means and removable concentric orifice plate means disposed within said circular opening; and

h) said removable orifice ring means and removable orifice plate means having within both a set of orifice openings which allow flow of reaction mixture from both the primary and secondary reservoirs to mix at right angles in close proximity to each other.

21. The apparatus of claim 20, wherein said removable orifice ring means and removable orifice plate means further comprise:

a) a three-layer orifice plate means having a lamination of two outer layers of a high rigidity material and a compressible elastomeric inner layer;

b) a means of applying a vertically oriented compressive force to said primary and secondary glass columns;

c) said vertically oriented compressive force being generated by an inner and an outer compression means positioned at a top of said glass columns in a terminating layer; and

d) said terminating layer also having a sealing means disposed between a plurality of drive fluid manifolds and said inner and an outer compression means to provide flow control.

22. The apparatus of claim 21, wherein said means of applying a vertically oriented compressive force comprises an inner and outer compression screw means positioned at the top of said primary and secondary glass columns in said terminating layer.

23. The apparatus of claim 22, wherein said sealing means comprises a compressible ferrule means.

24. The apparatus of claim 23, wherein said secondary reservoirs have air or other gases within an inlet chamber of the same secondary reservoirs, and said air or other gases are derived from a storage tank means which is connectively attached to said horizontal inlet channels of said secondary reservoirs, and wherein the pressure of the air or other gases within the inlet chamber of same said secondary reservoirs is about equal to a hydraulic pressure of said reaction mix in said horizontal outlet channel.

25. The apparatus of claim 24, wherein said temperature controlling means comprising a front and a back platen positioned so that they enclose an upper and lower belt, and said platens positioned so that they support said upper and lower belts to provide lateral deflection resistance to a pressurized molding space, said platens having a necessary rigidity and/or UV transparency and providing for the control of reaction temperatures in the molding space.

26. The apparatus of claim 25, wherein said platens are further comprised of:

a) a plurality of plates constructed of rigid thermally conductive and relatively low expansion materials selected from aluminum, stainless steel, Pyrex glass, ceramics or other thermally conductive materials;

b) said plates are arranged to provide a heating/cooling manifold with horizontal channels allowing for a flow of a coolant, and a gasket or spacer between said plates is provided to separate these channels;

c) a vertical hot and cold coolant supply manifold positioned such that one manifold is in each of the said plates comprising the platen and a single return line in said plate that is not adjacent to said molding space;

d) at an opening between each horizontal channel and each vertical supply manifold is a manually adjustable blending valve that can be set to vary the temperature of the coolant by blending variable amounts of hot and cold coolant entering the horizontal channel;

e) said platens are fixed in place by at least one support bracket on each side, and said platens are affixed to these supports at a minimum of at least 4 locations with finely adjustable screws that allow for alignment and adjustment of a gap between the front and back platens; and

f) each platen is provided with a means of connection on the lateral ends of said platen so that in can connectively attach to one or more additional platens as needed to provide the length required for a gelation time of the reaction mixture in conjunction with a belt speed.

27. The apparatus of claim 26, wherein said means for removing the polymerized gel from the mold space comprises an apparatus which unseals the lateral sides of an upper and lower belt, and then gels are cut and where supported gels are stacked by an actuated sideward shearing movement of a soft elastomeric platen located within said removal means.

28. The method of continuous casting of acrylamide or other gels for use in separations of proteins, nucleic acids or other biological materials comprising:

a) introducing into a molding space a reaction mixture comprising an reaction mixture capable of becoming a gel and initiators of its polymerization in a controlled manner, and wherein a casting manifold encloses said molding space and said casting manifold creates a continuously moving, sealed laminar cavity for maintaining a reacting gel solution at a modest pressure during a gelation process;

b) formulating a vertical gradient of composition in a plurality of components within said reaction mixture as said reaction mixture enters said molding space by pre-formulating the composition of an incremental reaction mix formula;

c) displacing air entrained in the reaction mixture within the molding space by use of a venting means disposed within said casting manifold;

d) initiating a polymerization process as said reaction mixture enters said mold space;

e) controlling a temperature of the reaction mixture during the gelation process as the reaction mixture travels through the apparatus;

f) removing a polymerized gel from the mold space, cutting the gel to a desired length, removing excess gel formed during the process, and stacking the polymerized, cut gels, further comprising;

1) a portion of the continuous gel moving into a cutting space in a cutting means;

2) said portion of the continuous gel is sheared from an upstream gel portion on a downstream side of a mold flash so that this flash will be attached to the downstream side of the next upstream gel, so that any mold flash, attached to the downstream side of the gel is cut off on the next round with this same shearing actuation at the downstream end of the gel;

3) a small extension of an actuating platen remains against the downstream shearing surface after a main platen returns in order to isolate the removed mold flash in a chamber at the extreme downstream end of the line where it can be washed away; and

4) stacking the completed gels or masks onto a washing belt or other support for further processing or stacking the completed gels or masks into running cassettes ready for use.