Non-specific binding resistant protein arrays and methods for making the same
First Claim
Patent Images
1. An array device comprising a substrate having a surface;
- and, one or more protein immobilization regions on said surface, said protein immobilization regions each comprising;
(i) an ordered hydrophobic monolayer formed of alkyl chains having proximal ends which are chemisorbed or physisorbed to said surface within said immobilization regions, and opposite hydrophobic distal ends;
(ii) a hydrophilic monolayer attached to said ordered hydrophobic monolayer, said hydrophilic monolayer comprising a set of first hydrophilic chains, each first hydrophilic chain having a proximal end by which said first hydrophilic chain is linked to an alkyl chain distal end, and an opposite hydrophilic distal end having a functional group for covalently attaching a protein capture agent thereto;
(iii) one or more of said protein capture agents attached to a first subset of said set of first hydrophilic chains within said immobilization regions through a residue formed by reacting said protein capture agents with said functional groups of said first subset of said first hydrophilic chains; and
, (iv) one or more second hydrophilic chains each being attached to a second subset of said set of said first hydrophilic chains through said functional group or residue thereof, wherein said second subset of first hydrophilic chains and said first subset of first hydrophilic chains are mutually exclusive of each other;
wherein said first hydrophilic chains and said second hydrophilic chains are effective to resist non-specific protein binding.
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Abstract
Arrays of protein-capture agents useful for the simultaneous detection of a plurality of proteins which are the expression products, or fragments thereof, of a cell or population of cells in an organism are provided. A variety of antibody arrays, in particular, are described. Methods of both making and using the arrays of protein-capture agents are also disclosed. The invention arrays are particularly useful for various proteomics applications including assessing patterns of protein expression and modification in cells.
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Citations
11 Claims
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1. An array device comprising
a substrate having a surface; - and,
one or more protein immobilization regions on said surface, said protein immobilization regions each comprising;
(i) an ordered hydrophobic monolayer formed of alkyl chains having proximal ends which are chemisorbed or physisorbed to said surface within said immobilization regions, and opposite hydrophobic distal ends;
(ii) a hydrophilic monolayer attached to said ordered hydrophobic monolayer, said hydrophilic monolayer comprising a set of first hydrophilic chains, each first hydrophilic chain having a proximal end by which said first hydrophilic chain is linked to an alkyl chain distal end, and an opposite hydrophilic distal end having a functional group for covalently attaching a protein capture agent thereto;
(iii) one or more of said protein capture agents attached to a first subset of said set of first hydrophilic chains within said immobilization regions through a residue formed by reacting said protein capture agents with said functional groups of said first subset of said first hydrophilic chains; and
,(iv) one or more second hydrophilic chains each being attached to a second subset of said set of said first hydrophilic chains through said functional group or residue thereof, wherein said second subset of first hydrophilic chains and said first subset of first hydrophilic chains are mutually exclusive of each other;
wherein said first hydrophilic chains and said second hydrophilic chains are effective to resist non-specific protein binding. - View Dependent Claims (2, 3, 4, 5)
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6. A method for making an array device comprising the steps of:
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a) providing a substrate having a surface; and
one or more protein immobilization regions on said surface, said protein immobilization regions each comprising;
i. an ordered hydrophobic monolayer formed of alkyl chains having proximal ends which are chemisorbed or physisorbed to said surface within said immobilization regions, and opposite hydrophobic distal ends;
ii. a hydrophilic monolayer attached to said ordered hydrophobic monolayer, said hydrophilic monolayer comprising a set of first hydrophilic chains, each first hydrophilic chain having a proximal end by which said first hydrophilic chain is linked to an alkyl chain distal end, and an opposite hydrophilic distal end;
iii. one or more functional groups, each for covalently attaching a protein capture agent thereto and each covalently attached to one or more of said hydrophilic distal ends of said first hydrophilic chains;
b) reacting one or more of said protein capture agents to one or more of said functional groups attached to a first subset of said set of first hydrophilic chains within said immobilization regions to attach said one or more protein capture agents through a residue formed by said reaction of said protein capture agents with said functional groups of said first subset of said first hydrophilic chains; and
,c) reacting one or more second hydrophilic chains with a second subset of said set of first hydrophilic chains so that one or more of said second hydrophilic chains attach through one or more residues of one or more of said second subset of said first hydrophilic chains, - View Dependent Claims (7, 8)
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9. A method for making an array device comprising the steps of:
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a) providing a substrate having a surface; and
one or more protein immobilization regions on said surface, said protein immobilization regions each comprising;
i. an ordered hydrophobic monolayer formed from a set of alkyl chains having proximal ends which are chemisorbed or physisorbed to said surface within said immobilization regions, and opposite hydrophobic distal ends;
ii. one or more functional groups for covalently attaching said protein captures agent thereto, each functional group being attached to said hydrophobic distal ends of a subset of said set of alkyl chains;
b) reacting one or more of said protein capture agents to one or more of said functional groups attached to a first subset of said set of alkyl chains within said immobilization regions to attach said one or more protein capture agents through a residue formed by said reaction of said protein capture agents with said functional groups of said first subset of said alkyl chains; and
,c) reacting one or more hydrophilic chains with a second subset of said set of alkyl chains so that one or more of said second hydrophilic chains attach through one or more residues of one or more of said second subset of said set of alkyl chains, wherein said first subset of said set of alkyl chains and said second subset of said set of alkyl chains are mutually exclusive. - View Dependent Claims (10, 11)
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Specification