Processing microtitre plates for covalent immobilization chemistries
Inventors
North, Stella H. • Lock, Evgeniya H. • Walton, Scott G. • Taitt, Chris Rowe
Assignees
Publication Number
US-8651158-B2
Publication Date
2014-02-18
Expiration Date
2030-11-17
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Abstract
Disclosed herein is a method of: treating an organic polymer with an electron beam-generated plasma; exposing the treated polymer to air or an oxygen- and hydrogen-containing gas, generating hydroxyl groups on the surface of the polymer; reacting the surface with an organosilane compound having a chloro, fluoro, or alkoxy group and a functional or reactive group that is less reactive with the surface than the chloro, fluoro, or alkoxy group; and covalently immobilizing a biomolecule to the functional or reactive group or a reaction product thereof.
Core Innovation
The invention provides a method for processing organic polymers, particularly polystyrene microtitre plates, to enable covalent immobilization of biomolecules. The method involves treating the polymer surface with an electron beam-generated plasma, exposing the treated polymer to air or an oxygen- and hydrogen-containing gas to generate hydroxyl groups on the surface, reacting the surface with an organosilane compound having a chloro, fluoro, or alkoxy group and a functional or reactive group less reactive with the surface than these groups, and covalently immobilizing a biomolecule to the functional or reactive group or its reaction product.
The problem addressed is the difficulty in controlling surface chemistry and achieving robust, functional covalent biomolecule attachment on common microtitre plate materials like polystyrene. Standard polystyrene plates rely on non-covalent adsorption, which can lead to protein denaturation, desorption, and loss of biomolecule activity. Existing covalent attachment approaches often depend on wet, corrosive chemical treatments that produce irregular surface etching and nonspecific attachment, or rely on deposited films that obscure the underlying polymer surface. Additionally, commercial covalent plates typically offer only a single attachment chemistry across all wells, limiting versatility.
To resolve these issues, the disclosed method uses a low-energy electron beam-generated plasma to selectively alter only the top few nanometers of the polymer surface, producing reactive hydroxyl groups without significant surface roughening or bulk property changes. This promotes uniform deposition of organosilane monolayers with functional groups oriented outward, improving covalent coupling efficiency and allowing multiple, distinct immobilization chemistries to be separately implemented within different wells of the same microtitre plate. This methodology combines dry plasma treatment with wet chemical silanization and crosslinking steps to create a customizable, multi-site bioassay platform that is cost-effective and adaptable to various biomolecules.
Claims Coverage
The patent contains one independent claim encompassing the main inventive features of the processing method and the articles produced.
Method for surface functionalization of an organic polymer
A method comprising treating an organic polymer with an electron beam-generated plasma and exposing the treated polymer to air or an oxygen- and hydrogen-containing gas to generate hydroxyl groups on the polymer surface.
Reaction of the polymer surface with an organosilane compound
Reacting the hydroxyl-containing polymer surface with an organosilane having a chloro, fluoro, or alkoxy group and a functional or reactive group less reactive with the surface than these groups.
Covalent immobilization of biomolecules
Covalently immobilizing a biomolecule to the functional or reactive group or a reaction product thereof on the polymer surface.
The claims cover a method of treating an organic polymer surface by electron beam plasma to generate hydroxyl groups, subsequent reaction with functional organosilanes, and covalent attachment of biomolecules, enabling multiple immobilization chemistries on polymer microtitre plates.
Stated Advantages
The plasma treatment produces a uniform, functionalized surface amenable to a broad range of chemistries with minimal surface roughening.
The method enables covalent immobilization chemistries that are tailorable to different biomolecules within discrete wells of a single microtitre plate.
Combining dry plasma treatment with wet silanization and crosslinking provides versatile, predictable functionality.
The approach reduces manufacturing costs compared to commercially available covalent attachment microplates.
The mild plasma treatment reduces hazardous chemical wastes and degradation relative to wet chemical methods.
Documented Applications
Surface modification of polystyrene microtitre plates for enzyme-linked immunosorbent assays (ELISAs) and other bioassays requiring covalent biomolecule immobilization.
Immobilization of peptides, antimicrobial peptides, antibodies, carbohydrates, lipids, and proteins for biomolecular interaction analyses.
Development of multi-site bioassay platforms allowing different covalent attachment chemistries in separate wells for high-throughput screening.
Tailoring microtitre plates for biomedical and biological applications including clinical diagnostics, monitoring of foodstuffs, vaccine and therapeutics development, and fundamental research.
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