Surface for the immobilization of ligands

Abstract

The present invention relates to binding surfaces for the immobilization of ligands, ligand surfaces and structured surface arrays which present a plurality of identical or different ligands. The invention further relates to a process for the production and the use of such surfaces and to specific binder molecules which can be used for the preparation thereof.

1 . Process for the generation of a binding layer on the metal surface of a solid phase carrier, comprising the step of contacting a solution of a plurality of identical or different anchor molecules represented by the formula  
       HS—R-M  
       wherein the structural moiety R provides for the formation of a self-assembling monolayer on the surface and M represents a mercaptophilic head group, with the surface under acidic conditions.  
     
     
         2 . Process according to  claim 1 , wherein a plurality of identical or different diluting molecules represented by the general formula  
       HS—R—X  
       wherein R is defined as in  claim 1  and X represents a non-mercaptophilic head group, is contacted with the surface together with the anchor molecules.  
     
     
         3 . Process according to  claim 2 , wherein the ratio of anchor molecules to diluting molecules ranges from 1:2 to 1:10.000.  
     
     
         4 . Binding surface comprising a binding layer obtainable according to the process of any of  claims 1  to  3  and a solid phase support.  
     
     
         5 . Binding surface according to  claim 4 , wherein a planar, non-structured carrier plate or a physically structured carrier plate having a plurality of separate fields for binding the anchors and diluants is used as the solid phase support.  
     
     
         6 . Measuring surface obtainable by covalently binding a plurality of identical or different biospecific interaction partners which carry a thiol group or which are modified by means of a thiol group to a binding surface according to any of claims  4  or  5 .  
     
     
         7 . Measuring surface according to  claim 6 , wherein the bound biospecific interaction partners are selected from proteins, peptides, oligonucleotides, carbohydrates, isoprenoids, enzymes, lipid structures, saccharides, antibodies, peptide hormones, cytokines, antibiotics and small organic molecules, the small organic molecules being characterized by having a molecular weight ranging from 50 to 3,000 g/mol.  
     
     
         8 . Measuring array comprising a plurality of measuring surfaces according to  claim 6  or  7 , wherein the individual measuring surfaces differ from each other at least in the structure of the bound interaction partners.  
     
     
         9 . Process for providing a measuring array according to  claim 8 , wherein solutions of the interaction partners are applied onto defined, spatially limited areas of the binding surface.  
     
     
         10 . Process according to  claim 9 , wherein the solution of the interaction partners is applied by means of a pipetting device, a spotting device, a micropipetting device or an ink-jet process.  
     
     
         11 . Process for the detection and/or quantification of an interaction between surface-bound and free interaction partners, comprising contacting the free interaction partners with a measuring surface or a measuring array according to any of  claims 6  to  8 .  
     
     
         12 . Process according to  claim 11 , wherein an interaction of the surface-bound interaction partners takes place with one or more receptors as free interaction partners, selected from proteins, DNA, RNA, oligonucleotides, prosthetic groups, vitamins, lipids, mono-, oligo- or polysaccharides or fusion proteins or synthetic primers.  
     
     
         13 . Use of a measuring surface or a measuring array according to any of  claims 6  to  8  for interaction analysis, in a screening process or in affinity chromatography.  
     
     
         14 . Use of a measuring surface or a measuring array according to any of  claims 6  to  8  in medical diagnostics.  
     
     
         15 . Anchor molecule of the formula  
       HS—R-M  
       wherein the structural moiety R provides for the formation of a self assembling to monolayer on the surface and M represents a mercaptophilic head group suitable for covalently binding to a thiol-functionalized ligand.  
     
     
         16 . Anchor molecule according to  claim 15 , wherein R comprises a hydrophobic structural moiety R a , formed by a branched or straight-chain hydrocarbon chain of 5 to 50 carbon atoms, which may be saturated or partially unsaturated.  
     
     
         17 . Anchor molecule according to  claim 15  or  16 , wherein R comprises a branched or non-branched hydrophilic spacer R b , formed by hydrocarbon chain of 5 to 1,000 carbon atoms which is interrupted by heteroatoms.  
     
     
         18 . Anchor molecule according to one of  claims 15  to  17 , wherein R has the following general formula  
       —(CH 2 ) a -Q 1 -(CH 2 ) b -{[Q 2 -(CH 2 ) c —[O—(CH 2 ) d ] e —O—(CH 2 ) f ] g -[Q 3 -(CH 2 ) c —[O—(CH 2 ) d′ ] e′ —O—(CH 2 ) f′ ] h } i -Q 4 -(CH 2 ) j -Q 5 -(CH 2 ) k — 
       wherein the variables are defined independently of each other as follows and the numeric ranges comprise their respective limiting values as well as the integers contained therein: 
 Q 1 ,Q 5  are —NH—C(O)—, —C(O)—NH— or a bond  
 Q 2 ,Q 3 ,Q 4  are —NH—C(O)— or —C(O)—NH— 
 a is 5 to 20, preferably 8 to 12, particularly preferred 10;  
 b is 0 to 5, preferably 0 if Q 1  is a bond and 1 to 10, preferably 2 to 7, particularly preferred 3 to 5in all other cases;  
 c, c′ are 1 to 5, preferably 1 to 3, particularly preferred 1;  
 d, d′ are 1 to 5, preferably 1 to 3, particularly preferred 2;  
 e, e′ are 1 to 5, preferably 1 to 3, particularly preferred 2;  
 f, f′ are 1 to 5, preferably 1 to 3, particularly preferred 1;  
 g, h are 0 to 3, provided that g+h≧1, preferably g+h=2;  
 i is 1 to 3, preferably 1 to 2, particularly preferred 1;  
 j is 0 to 5, preferably 1 to 3, particularly preferred 2; and  
 k is 0 to 5.  
 
     
     
         19 . Anchor molecule according to any of  claims 15  to  18 , wherein M is selected from iodo- or bromoacetamide, pyridylthio compounds, Michael acceptors, acrylic acid, (meth)acrylic acid esters, -amides, -lactones, -lactames, methylene-gem-difluorocyclopropanes, α,β-unsaturated aldehydes and ketones and α,β-unsaturated sulfones and sulfonamides.  
     
     
         20 . Anchor molecule according to one of  claims 15  to  18 , wherein M has the following structure  
       
         
           
           
               
               
           
         
       
       wherein 
 R 1  and R 2  are independently hydrogen or C 1 -C 5  alkyl,  
 R 3  and R 4  are independently hydrogen or C 1 -C 5  alkyl or  
 R 3  and R 4 together form ═O.  
 
     
     
         21 . Process for the solid phase synthesis of an anchor molecule according to any of  claims 15  to  20 , wherein the anchor molecule is coupled to the synthesis phase via the thiol group and is cleaved off in an acidic medium to provide the free anchor molecule.