Irregularly-shaped macroporous copolymer particles and methods of using same
A matrix comprising irregularly-shaped macroporous copolymer particles is disclosed. The matrix can be employed liquid chromatography applications, such as detecting the presence or absence of a heteroduplex structure in a mixture of hetero- and homoduplex structures. The non-monolithic crushed macroporous copolymer network can be packed in a chromatography column, and such columns facilitate high resolution separations, while maintaining low back pressures, short separation times and long column lifetimes. The irregularly-shaped macroporous copolymer particles can also be employed in analyte isolation operations.
1 . A liquid chromatography column comprising:
(a) a matrix adapted for use in a chromatography technique selected from the group consisting of denaturing high performance liquid chromatography, high performance liquid chromatography and reversed phase ion paring chromatography, the matrix comprising irregularly-shaped macroporous copolymer particles; and
(b) a durable support structure.
2 . The liquid chromatography column of claim 1 , wherein the durable support structure is selected from the group consisting of a stainless steel tube, a PEEK™ tube and an HDPE tube, a glass tube and a titanium steel tube.
3 . The liquid chromatography column of claim 1 , wherein the irregularly-shaped macroporous copolymer particles are formed by:
(a) stirring a polymerization mixture comprising a monomer and a crosslinker; and
(b) while stirring, employing a polymerization technique selected from the group consisting of chain-growth polymerizations and step-growth polymerizations to form irregular-shaped macroporous copolymeric particles.
4 . The liquid chromatography column of claim 3 , wherein the monomer comprises a molecule comprising a polymerizable vinyl group.
5 . The liquid chromatography column of claim 3 , wherein the chain polymerization comprises free radical-initiated polymerization
6 . The liquid chromatography column of claim 3 , wherein the step polymerization comprises condensation polymerization.
7 . The liquid chromatography column of claim 1 , wherein the irregularly-shaped macroporous copolymer particles are formed by:
(a) polymerizing a monomer to form a structurally rigid material by employing a polymerization technique selected from the group consisting of chain-growth polymerizations and step-growth polymerizations; and
(b) disrupting the structural integrity of the structurally rigid material to form a powder.
8 . The liquid chromatography column of claim 7 , wherein the monomer comprises a molecule comprising a polymerizable vinyl group.
9 . The liquid chromatography column of claim 7 , wherein the chain polymerization comprises free radical-initiated polymerization.
10 . The liquid chromatography column of claim 7 , wherein the step polymerization comprises condensation polymerization.
11 . A liquid chromatography method of isolating an analyte from a sample known or suspected to comprise an analyte, the method comprising:
(a) contacting a sample known or suspected to comprise an analyte with a matrix comprising. irregularly-shaped macroporous copolymer particles; and
(b) isolating the analyte from the matrix by employing a technique selected from the group consisting of HPLC, DHPLC and RPIPC.
12 . The method of claim 11 , wherein the analyte is selected from the group consisting of an organic molecule, an inorganic molecule, a bioinorganic molecule, an ion and a biomolecule.
13 . The method of claim 12 , wherein the organic molecule is a polymer.
14 . The method of claim 12 , wherein the biomolecule is selected from the group consisting of a nucleic acid, a nucleic acid oligomer, a peptide and a protein.
15 . The method of claim 14 , wherein the nucleic acid oligomers are selected from the group consisting of DNA, mutant DNA, RNA, and mutant RNA.
16 . The method of claim 11 , wherein the method is automated.
17 . A method of separating a mixture comprising double stranded DNA segments of different lengths, the method comprising:
(a) contacting a mixture known or suspected to comprise double stranded DNA segments of different lengths with an ion pairing agent to form an ion paired species;
(b) contacting the ion paired species with a matrix comprising irregularly-shaped macroporous copolymer particles; and
(c) eluting the double stranded DNA segments with an elution solvent.
18 . The method of claim 17 , wherein the ion pairing agent is selected from the group consisting of alkylammonium salts of organic acids and alkylammonium salts of inorganic acids.
19 . The method of claim 17 , wherein the elution solvent is selected from the group consisting of water, an organic solvent and an aqueous buffer.
20 . The method of claim 19 , wherein the organic solvent is selected from the group consisting of organic solvents mixable with water, acetonitrile, methanol, and THF.
21 . The method of claim 17 , wherein the method is automated.
22 . A method of separating a homoduplex nucleic acid structure from a heteroduplex nucleic acid structure, the method comprising:
(a) providing a mixture known or suspected to comprise at least one structure selected from the group consisting of a homoduplex nucleic acid structure and a heteroduplex nucleic acid structure, the mixture further comprising an ion-paring reagent;
(b) contacting the mixture with a matrix comprising irregularly-shaped macroporous copolymer particles under conditions known or suspected to partially denature the at least one structure;
(c) eluting the nucleic acid structures with an elution solvent, whereby a homoduplex nucleic acid structure is separated from a heteroduplex nucleic acid structure.
23 . The method of claim 22 , wherein the ion pairing reagent is selected from the group consisting of an alkylammonium salt of an organic acid and an alkylammonium salt of an inorganic acid.
24 . The method of claim 22 , wherein the elution solvent is selected from the group consisting of water, an organic solvent and an aqueous buffer.
25 . The method of claim 24 , wherein the organic solvent is selected from the group consisting of acetonitrile and methanol.
26 . The method of claim 22 , wherein the method is automated.
27 . The method of claim 22 , wherein the method is repeated a desired number of times.