Hydrogels and hydrogel arrays made from reactive prepolymers crosslinked by [2 + 2] cycloaddition

Abstract

Reactive prepolymers incorporating [2+2] photoreactive sites are synthesized. Upon exposure to UV light, these prepolymers undergo [2+2] cycloaddition to crosslink. When crosslinked, the reactive prepolymers form a hydrogel. Selective hydrogel formation is provided through selective exposure of the reactive prepolymer to UV light. Supports and other molecules may be attached or incorporated into the hydrogel through [2+2] cycloaddition with uncrosslinked [2+2] photoreactive sites present in the hydrogel.

Description

REFERENCE TO RELATED APPLICATIONS[0001] This application is a continuation-in-part of U.S. Nonprovisional Application No. 09 / 344,217, filed Jun. 25, 1999, entitled "Polyacrylamide Hydrogels and Hydrogel Arrays Made from Polyacrylamide Reactive Prepolymers," which claimed the benefit of U.S. Provisional Application No. 60 / 109,821, filed Nov. 25, 1998 entitled "Polyacrylamide Hydrogels and Hydrogel Arrays Made from Polyacrylamide Reactive Prepolymers."[0002] Acrylamide (CH.sub.2.dbd.CHC(O)NH.sub.2; C.A.S. 79-06-1; also known as acrylamide monomer, acrylic amide, propenamide, and 2-propenamide) is an odorless, free-flowing white crystalline substance that may be polymerized to form polyacrylamides. The resulting high molecular weight polymers have a variety of uses and further can be modified to optimize nonionic, anionic, or cationic properties for specified uses.[0003] Polyacrylamide hydrogels are used as molecular sieves for the separation of nucleic acids, proteins, peptides, oligo...

AI Technical Summary

Benefits of technology

[0010] Polyacrylamide hydrogels that incorporate [2+2] photoreactive sites are disclosed. The hydrogels are especially useful for microarray formation and are made from prepolymers, including polyacrylamide reactive prepolymers. The photoreactive sites allow use of [2+2] cycloaddition reactions to not only crosslink the polyacrylamide reactive prepolymers forming the hydrogel, but also for later attachment of any other molecules incorporating additional photoreactive sites. The disclosed hydrogels provide a more uniform pore size, likely resulting from an improved control of crosslinking, making them preferable for use with DNA based probes. Additionally, because higher viscosity prepolymers, as opposed to low viscosity monomer solutions, are used to form the arrays, manufacturing is simplified.

[0054] Crosslinking of the [2+2] photoreactive sites of the photocyclizable monomers in the reactive prepolymer is most preferably done with ultraviolet irradiation. Optionally, a photosensitiser may be added to the reactive prepolymer to increase the efficiency of the cycloaddition reaction. Preferred photosensitisers include water-soluble quinones and xanthones, including anthroquinone, thioxanthone, sulfonic acid quinone, benzoin ethers, acetophenones, benzoyl oximes, acylphosphines, benzophenones, and TEMED (N,N,N',N'-tetramethylethylendia-mine). Anthroquinone-2-sulfonic acid is most preferred and is available from ALDRICH, Milwaukee, Wis.

[0073] While not necessary, hydrogels may also be formed by adding an additional crosslinking agent to the reactive prepolymer before UV irradiation. The inclusion of an additional crosslinking agent increases the amount of crosslinking between the reactive prepolymers. Preferable additional crosslinking agents include pentaerythritol tetraacrylate. Additionally, crosslinking agents can be [2+2] cyclized with acrylate based reactive prepolymers to form hydrogels. Thus, while acrylate based reactive prepolymers will not cyclize with themselves, they can be cyclized into hydrogels through the addition of an additional crosslinking agent.

Problems solved by technology

1. The coating process is difficult and expensive to automate because the film thickness is controlled with a spacer and top glass plate. The removal of the top glass plate must be done manually. Due to difficulties automating the process (e.g., viscosity too low for commercial coating methods), the coating of the monomer solution currently is done manually.

2. The reaction time of the acrylamide is excessively long (e.g., typically from about 15 to about 90 minutes at a short wavelength of about 254 nm), making the UV polymerization and crosslinking step incompatible with standard imaging equipment such as mask aligners and photoprinters.

3. The acrylamide monomer is a neurotoxin and a carcinogen which makes coating, handling, and waste disposal of the material hazardous and expensive.

Added to these disadvantages are the further problems that crystallization of monomer frequently occurs on commonly used equipment and laboratory surfaces, and exothermic polymerization can occur in coating reservoirs, necessitating the use of stabilizers or inhibitors.

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