Cast-to-shape electrokinetic trapping medium
View Patent ↗A three-dimensional microporous polymer network material, or monolith, cast-to-shape in a microchannel. The polymer monolith, produced by a phase separation process, is capable of trapping and retaining charged protein species from a mixture of charged and uncharged species under the influence of an applied electric field. The retained charged protein species are released from the porous polymer monolith by a pressure driven flow in the substantial absence of the electric field. The pressure driven flow is independent of direction and thus neither means to reverse fluid flow nor a multi-directional flow field is required, a single flow through the porous polymer monolith can be employed, in contrast to prior art systems. The monolithic polymer material produced by the invention can function as a chromatographic medium. Moreover, by virtue of its ability to retain charged protein species and quantitatively release the retained species the porous polymer monolith can serve as a means for concentrating charged protein species from, for example, a dilute solution.
1. A method for controllably retaining and releasing charged species contained in solution, comprising:
a) passing the solution through a porous stationary phase disposed in a capillary column by imposing a voltage gradient between first and second ends of the column; and
b) subsequently subjecting the capillary column to a pressure gradient between first and second ends of the column, wherein the porous stationary phase is comprised of a porous polymer monolith prepared by the method of;
pretreating the internal surface of the capillary column;
filling the capillary column with a solution comprising a monomer mixture and a solvent, wherein the monomer mixture comprises an alkoxy protected metal functionality, a crosslinker, and a polymerization initiator;
polymerizing the monomer by exposing at least a portion of the mixture to radiation to form a porous polymer monolith;
hydrolyzing the alkoxy functionalities by washing the polymer monolith with a hydrolyzing solution; and
washing the hydrolyzing solution from the polymer monolith.
2. The method of claim 1 , wherein said step of pretreating includes exposing the internal surface to an aqueous solution of acetic acid.
3. The method of claim 1 , wherein the monomer mixture is a mixture of lauryl acetate, 1,6-hexyldiol diacrylate, and 3-(trimethoxy silyl)propylacrylate.
4. The method of claim 1 , wherein the metal comprising the alkoxy protected metal functionality is Si, Ti or Zr.
5. The method of claim 1 , wherein said step of polymerizing includes exposing the monomer solution to UV radiation.
6. The method of claim 1 , wherein said step of hydrolyzing includes exposing the polymer monolith to a solution of acetic acid in acetonitrile.
7. The method of claim 1 , further including the step of adding a polymerization inhibitor to improve resolution between irradiated and unirradiated regions of the monomer mixture.
8. The method of claim 1 , further including the step of purging the solvent from the monolith prior to the step of hydrolysis.
9. The method of claim 1 , wherein said step of pretreating includes exposing the internal surface to an aqueous solution of acetic acid and 3-(trimethoxy silyl)propylacrylate.
10. The method of claim 8 , wherein said step of purging includes rinsing the polymer monolith with 2-methoxyethanol.
11. The method of claim 1 wherein at least 25 vol % of the monomer contains a hydrophobic group such as alkyl, aryl, or substituted versions thereof.
12. A method for treating a solution to separate charged species from uncharged species contained therein, comprising:
a) passing the solution through a porous stationary phase disposed in a column by imposing a voltage gradient along the column, wherein the porous stationary phase is a porous polymer monolith produced by the method of claim 1 ; and
b) subsequently subjecting the column to a pressure differential.
13. A method for chromatographic separation of protein species, comprising:
passing a solution containing species to be separated through a porous stationary phase disposed in a column by imposing a voltage gradient along the column, where in the porous stationary phase is produced by the method of claim 1 .