Attachment of molecules to surfaces
View Patent ↗The present invention relates to methods, reagents, and substrates that can be used for, for example, immobilizing biomolecules, such as nucleic acids and proteins. In an embodiment, the present invention relates to surfaces coated with a polymer according to the present invention. In an embodiment, the present invention relates to methods for thermochemically and/or photochemically attaching molecules to a surface at a high density.
1. A method of attaching biomolecule to surface of substrate, the method comprising:
providing composition comprising polymer;
the polymer comprising:
at least about 40 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof in an amount effective to provide a contact angle of about 30° to about 60° for a sessile water drop; and
one or more pendent reactive groups configured to form covalent bonds with corresponding functional groups on biomolecule; wherein the one or more pendent reactive groups are selected from the group consisting of thermochemically reactive groups, photoreactive groups, or mixtures thereof;
the polymer being configured to be covalently attached to the surface coating and immobilizing the composition on the substrate surface;
providing solution comprising biomolecule comprising one or more functional groups reactive with the reactive groups;
applying an aliquot of the solution to the substrate surface; and
forming covalent bonds between the reactive group and the functional group of the biomolecule.
2. The method of claim 1 , wherein the polymer comprises N,N -dimethylacrylamide, N,N-diethylacrylamide, N-isopropylacrylamide, N-t-butylacrylamide, N-octylacrylamide, N-cyclohexylacrylamide, N-phenylacrylamide, N-benzylacrylamide, N—(CH 2 OC 4 H 9 )acrylamide, N—(CH 2 CH 2 OH)acrylamide, N—(CH 2 OH)acrylamide, N—(CH 2 CH 2 CH 2 OH)acrylamide, N-methylmethacrylamide, N-ethylmethacrylamide, N,N-dimethylmethacrylamide, N,N-diethylmethacrylamide, N-acryloylmorpholine, N-methacryloylmorpholine, N—(CH 2 OC 4 H 9 )methacrylamide, N—(CH 2 CH 2 OH)methacrylamide, N—(CH 2 OH)methacrylamide, or mixtures thereof.
3. The method of claim 2 , wherein the polymer comprises N,N-dimethylacrylamide.
4. The method of claim 1 , wherein the polymer comprises about 85 to about 95 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof.
5. A method of attaching biomolecule to surface of substrate, the method comprising:
providing composition comprising polymer;
the polymer comprising:
at least about 40 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof; and
one or more pendent reactive groups configured to form covalent bonds with corresponding functional groups on biomolecule; wherein the one or more reactive groups comprise thermochemically reactive group; the thermochemically reactive group comprising NOS, epoxide, aldehyde, isothiocyanate, or mixtures thereof;
the polymer being configured to be covalently attached to the surface coating and immobilizing the composition on the substrate surface;
providing solution comprising biomolecule comprising one or more functional groups reactive with the reactive groups;
applying an aliquot of the solution to the substrate surface; and
forming covalent bonds between the reactive group and the functional group of the biomolecule.
6. A method of attaching biomolecule to surface of substrate, the method comprising:
providing composition comprising polymer;
the polymer comprising:
at least about 40 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof; and
one or more pendent reactive groups configured to form covalent bonds with corresponding functional groups on biomolecule; wherein the one or more reactive groups comprise photochemically reactive group; the photochemically reactive group comprising acetophenone, benzophenone, quinone, anthraquinone, anthrone, heterocyclic analog of anthrone, or mixtures thereof;
the polymer being configured to be covalently attached to the surface coating and immobilizing the composition on the substrate surface;
providing solution comprising biomolecule comprising one or more functional groups reactive with the reactive groups;
applying an aliquot of the solution to the substrate surface; and
forming covalent bonds between the reactive group and the functional group of the biomolecule.
7. The method of claim 6 , wherein the one or more reactive groups comprise photochemically reactive group; the photochemically reactive group comprises photoreactive aryl ketone; and the biomolecule comprises nucleic acid.
8. The method of claim 1 , wherein the substrate comprises activated slide.
9. The method of claim 8 , wherein the slide is configured for fabricating microarray; the biomolecule comprises nucleic acid; and the surface comprises the surface of plastic, silicon hydride, or organosilane-pretreated glass or silicon slide.
10. The method of claim 8 , wherein the slide is configured to receive sample in an amount of twenty nanoliters or less.
11. The method of claim 1 , wherein the biomolecule comprises polypeptide or nucleic acid.
12. The method of claim 11 , wherein the biomolecule comprises nucleic acid and the nucleic acid comprises amine group, sulfhydryl group, or mixture thereof.
13. The method of claim 5 , wherein the polymer comprises N,N-dimethylacrylamide, N,N-diethylacrylamide, N-isopropylacrylamide, N-t-butylacrylamide, N-octylacrylamide, N-cyclohexylacrylamide, N-phenylacrylamide, N-benzylacrylamide, N—(CH 2 OC 4 H 9 )acrylamide, N—(CH 2 CH 2 OH)acrylamide, N—(CH 2 OH)acrylamide, N—(CH 2 CH 2 CH 2 OH)acrylamide, N-methylmethacrylamide, N-ethylmethacrylamide, N,N-dimethylmethacrylamide, N,N-diethylmethacrylamide, N-acryloylmorpholine, N-methacryloylmorpholine, N—(CH 2 OC 4 H 9 )methacrylamide, N—(CH 2 CH 2 OH)methacrylamide, N—(CH 2 OH)methacrylamide, or mixtures thereof.
14. The method of claim 5 , wherein the polymer comprises N,N -dimethylacrylamide.
15. The method of claim 5 , wherein the polymer comprises about 85 to about 95 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof.
16. The method of claim 6 , wherein the polymer comprises N,N -dimethylacrylamide, N,N-diethylacrylamide, N-isopropylacrylamide, N-t-butylacrylamide, N -octylacrylamide, N-cyclohexylacrylamide, N-phenylacrylamide, N-benzylacrylamide, N—(CH 2 OC 4 H 9 )acrylamide, N—(CH 2 CH 2 OH)acrylamide, N—(CH 2 OH)acrylamide, N—(CH 2 CH 2 CH 2 OH)acrylamide, N-methylmethacrylamide, N-ethylmethacrylamide, N,N -dimethylmethacrylamide, N,N-diethylmethacrylamide, N-acryloylmorpholine, N -methacryloylmorpholine, N—(CH 2 OC 4 H 9 )methacrylamide, N—(CH 2 CH 2 OH)methacrylamide, N—(CH 2 OH)methacrylamide, or mixtures thereof.
17. The method of claim 6 , wherein the polymer comprises N,N -dimethylacrylamide.
18. The method of claim 6 , wherein the polymer comprises about 85 to about 95 mol-% N-substituted acrylamide, N,N-disubstituted acrylamide, N-substituted methacrylamide, N,N-disubstituted methacrylamide, or mixture thereof.