Biomolecule retaining material and methods for attaching biomolecules to a surface

Abstract

The present invention provides a method and material for attachment of biomolecules onto the surface of a substrate, such as microwell plates, tubes, beads, microscope slides, silicon wafers or membranes. The material includes a substrate having a surface coating including a polyamine compound. In one embodiment, the method and material are used to immobilize nucleic acid probes onto plastic materials such as microwell plates, e.g., for use in hybridization assays. In particular, the material can be used to attach a biomolecule (e.g., a nucleic acid) which in turn can be used for specific binding reactions (e.g., to hybridize a nucleic acid to its complementary strand).

What is claimed is:  
     
         1 . A biomolecule retaining material, comprising a substrate having a surface coating comprising a polyamine compound on at least a portion of a surface of the substrate.  
     
     
         2 . The material of  claim 1 , further comprising a biomolecule non-covalently bound to said polyamine compound.  
     
     
         3 . The material of  claim 2 , wherein said biomolecule is DNA.  
     
     
         4 . The material of  claim 1 , wherein said polyamine compound comprises a diamine.  
     
     
         5 . The material of  claim 1 , wherein said polyamine compound comprises a triamine.  
     
     
         6 . The material of  claim 1 , wherein said polyamine compound comprises (OR 1 ) 3 S 1 —CH 2 (CH 2 CH 2 NH) n —H, wherein n is an integer of two or greater and R 1  is hydrogen or a lower alkyl having from one to four carbon atoms.  
     
     
         7 . The material of  claim 1 , wherein said polyamine compound comprises (OR 1 ) 3 S 1 —CH 2 (CH 2 CH 2 NR 2 )—H, wherein n is an integer of two or greater, R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, and R 2  is an alkyl, acyl, anhydride, or ester.  
     
     
         8 . The material of  claim 1 , wherein said polyamine compound comprises trialkoxysilylpropyl ethylenediamine.  
     
     
         9 . The material of  claim 1 , wherein said polyamine compound comprises trialkoxysilylpropyl diethylenetriamine.  
     
     
         10 . The material of  claim 1 , wherein said polyamine compound comprises (OR 1 ) 3 Si—(CH 2 ) n —R 2 —(CH 2 ) m —NH—(CH 2 ) p NR 3 H, wherein R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, R 2  is a methylene group or a conjugated group, R 3  is hydrogen or a conjugated group, provided that at least one of R 2  or R 3  is a conjugated group, n is an integer of one or greater, m is an integer of one or greater, and p is an integer of one or greater.  
     
     
         11 . The material of  claim 10 , wherein at least one of R 2  or R 3  is an aromatic group or a conjugated diene.  
     
     
         12 . The material of  claim 10 , wherein said polyamine compound comprises (aminoethylaminomethyl)phenethyl trimethoxysilane.  
     
     
         13 . The material of  claim 10 , wherein said polyamine compound comprises 3-(N-styrylmethyl-2-amino ethyl amino)propyltrimethoxysilane.  
     
     
         14 . The material of  claim 1 , wherein said polyamine compound comprises (OR 1 ) 3 Si—(CH 2 ) n —NH—(CH 2 ) m —NH 2 , wherein R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, n is an integer of one or greater, and m is an integer of one or greater.  
     
     
         15 . The material of  claim 14 , wherein said polyamine compound comprises N-(6-aminohexyl)aminopropyl trimethoxysilane.  
     
     
         16 . The material of  claim 1 , wherein at least one amine is alkylated.  
     
     
         17 . The material of  claim 1 , wherein at least one amine is over-alkylated.  
     
     
         18 . The material of  claim 1 , wherein at least one amine is reacted to prevent any subsequent bond formation with a biomolecule.  
     
     
         19 . The material of  claim 1 , wherein said substrate is glass.  
     
     
         20 . A method for treating a biomolecule retaining material, comprising coating at least a portion of a substrate with a polyamine compound and treating the polyamine compound with an end capping reagent suitable to increase the water contact angle of the coated substrate.  
     
     
         21 . The method of  claim 20 , wherein said end capping reagent comprises an alkyl halide, acyl halide, anhydride, ester, or combinations thereof.  
     
     
         22 . The method of  claim 20 , wherein said treating results in a coated substrate having a water contact angle greater than about 46°.  
     
     
         23 . The method of  claim 20 , wherein said treating results in a coated substrate having a water contact angle greater than about 70°.  
     
     
         24 . The method of  claim 20 , wherein said treating results in a coated substrate having a water contact angle of from about 50° to about 80°.  
     
     
         25 . The method of  claim 20 , wherein said polyamine compound comprises a diamine.  
     
     
         26 . The method of  claim 20 , wherein said polyamine compound comprises a triamine.  
     
     
         27 . The method of  claim 20 , wherein said polyamine compound comprises (OR 1 ) 3 S 1 —CH 2 (CH 2 CH 2 NH) n —H, wherein n is an integer of two or greater and R 1  is hydrogen or a lower alkyl having from one to four carbon atoms.  
     
     
         28 . The method of  claim 20 , wherein said polyamine compound comprises (OR 1 ) 3 S 1 —CH 2 (CH 2 CH 2 NH)) n —H, wherein n is an integer of two or greater, R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, and R is an alkyl, acyl, anhydride, or ester.  
     
     
         29 . The method of  claim 20 , wherein said polyamine compound comprises trialkoxysilylpropyl ethylenediamine.  
     
     
         30 . The method of  claim 20 , wherein said polyamine compound comprises trialkoxysilylpropyl diethylenetriamine.  
     
     
         31 . The method of  claim 20 , wherein said polyamine compound comprises (OR 1 ) 3 Si—(CH 2 ) n —R 2 —(CH 2 ) m —NH—(CH 2 ) p NR 3 H, wherein R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, R 2  is a methylene group or a conjugated group, R 3  is hydrogen or a conjugated group, provided that at least one of R 2  or R 3  is a conjugated group, n is an integer of one or greater, m is an integer of one or greater, and p is an integer of one or greater.  
     
     
         32 . The method of  claim 31 , wherein at least one of R 2  or R 3  is an aromatic group or a conjugated diene.  
     
     
         33 . The method of  claim 31 , wherein said polyamine compound comprises (aminoethylaminomethyl)phenethyl trimethoxysilane.  
     
     
         34 . The method of  claim 31 , wherein said polyamine compound comprises 3-(N-styrylmethyl-2-aminoethylamino)propyltrimethoxysilane.  
     
     
         35 . The method of  claim 20 , wherein said polyamine compound comprises (OR 1 ) 3 Si—(CH 2 ) n —NH—(CH 2 ) m —NH 2 , wherein R 1  is hydrogen or a lower alkyl having from one to four carbon atoms, n is an integer of one or greater, and m is an integer of one or greater.  
     
     
         36 . The method of  claim 35 , wherein said polyamine compound comprises N-(6-aminohexyl)aminopropyl trimethoxysilane.  
     
     
         37 . The method of  claim 20 , further comprising alkylating at least one amine.  
     
     
         38 . The method of  claim 20 , further comprising over-alkylating at least one amine.  
     
     
         39 . The method of  claim 20 , further comprising reacting at least one amine to prevent any subsequent bond formation with a biomolecule.  
     
     
         40 . The method of  claim 20 , wherein said substrate is glass.  
     
     
         41 . The method of  claim 20 , further comprising non-covalently binding a biomolecule to said coated substrate.  
     
     
         42 . The method of  claim 41 , wherein said biomolecule is DNA.