Polymer brushes for immobilizing molecules to a surface and having water-soluble or water-dispersible segments therein and probes bonded thereto

• US 6,833,276 B2
• Filed: 07/24/2001
• Issued: 12/21/2004
• Est. Priority Date: 07/02/1999
• Status: Expired due to Fees

• Current Assignee: Freeslate Inc. (Unchained Labs Inc.)
• Original Assignee: Symyx Technologies Incorporated (Dassault Systemes SA)
• Inventors: Charmot, Dominique, Klaerner, Gerrit, Jandeleit, Bernd, Mansky, Paul, Mazzola, Laura T., Benoit, Didier, Nielsen, Ralph B.
• Primary Examiner(s): CEPERLEY, MARY
• Application Number:
  US09/911,683
• Publication Number:
  US 20020001845A1
• Time in Patent Office: 1,246 Days
• Field of Search: 436/532, 436/531, 436/527, 436/546, 435/6, 530/408, 530/409, 530/391.1
• US Class Current: 436/532
• CPC Class Codes: C08F 220/54 Amides , e.g. N,N-dimethylacrylamide or N-isopropylacrylamide
• CPC Class Codes: C08F 4/00 Polymerisation catalysts
• CPC Class Codes: G01N 27/44747 Composition of gel or of carrier mixture
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: Y10T 428/8305 Miscellaneous [e.g., treated surfaces, etc.]
• CPC Class Codes: Y10T 436/10 Composition for standardization, calibration, simulation, stabilization, preparation or preservation; processes of use in preparation for chemical testing

Abstract

Sensors for determining the presence and concentration of bio-molecules in a biological sample are provided in the form of polymer brushes, which comprise a substrate having a surface that is modified with a water-dispersible or water-soluble polymer segment having functional groups that bind probes. The method of synthesis of such sensors preferably includes use of controlled free radical polymerization techniques, and in particular the use of an iniferter initiator, which allows for controlled architecture polymers to modify the surface of the substrate. In this manner functional groups in the polymer chain are removed from the surface, which allows for solution chemistry to be more realistically reproduced with the benefits of a solid bound probe.

Claims

1. A method of preparing a sensor for detecting a biological molecule in an aqueous sample, the method comprising:

bonding an iniferter initiator to a substrate surface at one or more points to form a derivatized surface, said surface-bound iniferter initiator having the formula;

wherein C is a moiety on the surface of the substrate;

L is a linker group capable of bonding to at least one C moiety;

q, r and t are independently 0 or 1, provided the sum of q+r+t is at least 1;

Y is a residue capable of initiating free radical polymerization upon homolytic cleavage of the Y—

S bond;

S is sulfur; and

, G is a nitrogen or an oxygen heteroatom;

contacting said derivatized surface with a composition comprising a water-soluble or water-dispersible free radically polymerizable monomer mixture under reaction conditions to form bound polymer chains comprising a water-dispersible segment having a weight average molecular weight of at least about 1000 grams per mole, wherein (i) the mixture comprises a monomer that has one or more functionalized sites thereon for reaction with a probe selective for the biological molecule and a monomer that does not have a functionalized site for reaction with said probe, and (ii) the mixture comprises an acrylamide-based monomer and at least 1 other monomer; and

bonding the probe to the bound polymer chains through the functionalized sites.

2. The method according to claim 1 wherein the surface bound iniferter initiator has the formula:

wherein N is nitrogen, and R¹ and R² are independently selected from hydrocarbyl and substituted hydrocarbyl.

3. The method according to claim 2 wherein said surface bound iniferter initiator has the formula:

wherein Et is ethyl.

4. The method according to claim 3 wherein C is derived from a hydroxyl group.

5. The method according to claim 3 wherein q=1, r=1 and t=0.

6. The method according to claim 3 wherein q=1, r=1 and t=1.

7. The method according to claim 1 further comprising bonding spacer molecules to said substrate surface at one or more points to form the derivatized surface, said one or more points being different from the points at which said bound polymer chains are formed to space said bound polymer chains apart from each other, wherein the ratio of polymer chains to the sum of polymer chains and spacer molecules is about 0.75:

1.

8. The method according to claim 1 wherein the acrylamide-based monomer has the formula:

wherein R⁴ is H or an alkyl group, R⁵ is methyl and R⁶ is methyl.

9. The method according to claim 8 wherein the monomer is N,N-dimethylacrylamide.

10. The method according to claim 1 further comprising separating unbound polymer after said derivatized surface is contacted with a composition comprising the monomer mixture to form polymer chains.

11. The method according to claim 10 wherein the monomer mixture additionally comprises an unbound iniferter initiator.

12. The method according to claim 1, wherein the monomer mixture comprises about 10% to about 90% of the monomer having said functionalized sites for reaction with said probe, based on the total mass of monomer in said mixture.

13. The method according to claim 1 wherein the monomer mixture comprises about 15% to about 50% of the monomer having said functionalized sites for reaction with said probe, based on the total mass of monomer in said mixture.

14. The method according to claim 1 wherein the one or more functionalized sites on said functionalized monomer are in their a active state for reaction with said probe.