Antibody-based protein array system
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Abstract
The invention of novel protein microarrays and protein microarray-based techniques to determine the presence and amounts of proteins of interest are described. These microarrays and methods of use can be used for the simultaneous detection of a multiplicity of antigens or antibodies in a high throughput assay based upon the differential affinity of molecules for one another. The microarrays can be formed by immobilizing capture proteins in an array on a membrane. Analytes of interest can be bound by the capture proteins and can be detected either by the position to which they are immobilized or by the identity of detecting proteins or agents which bind to the analytes of interest. The interactions that can be detected using the present invention can also be used to characterize proteins of unknown identity or character.
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Citations
95 Claims
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1-63. -63. (canceled)
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64. A substrate for detecting one or more cytokines in a sample comprising:
a microarray of capture antibodies immobilized on a membrane, wherein the microarray comprises an ordered, two-dimensional arrangement of at least 10 array elements, wherein each array element comprises at least one capture antibody specific for a certain type of cytokine, and wherein at least two array elements are specific for different types of cytokines. - View Dependent Claims (65, 66, 67)
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68. A method for detecting one or more cytokines in a sample comprising:
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a) passing a solution containing one or more types of cytokines over a microarray of capture antibodies immobilized on a membrane, wherein the microarray comprises an ordered, two-dimensional arrangement of at least 10 array elements, wherein each array element comprises at least one capture antibody specific for a certain type of cytokine, and wherein at least two array elements are specific for different types of cytokines, wherein at least one type of cytokine in the solution binds to its respective capture antibody on the membrane;
b) adding a solution of one or more types of labeled detection antibodies to the membrane, wherein at least one type of labeled detection antibody in the solution is capable of binding the bound cytokine; and
c) detecting bound labeled detection antibody, the detection of bound labeled detection antibody in a position on the microarray corresponding to an array element having a capture antibody specific for a certain type of cytokine indicating the presence of that type of cytokine. - View Dependent Claims (69, 70, 71)
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72. A method for detecting one or more specified proteins in a sample comprising:
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a) passing a solution containing one or more types of specified protein over a microarray of capture proteins immobilized on a membrane, wherein the microarray comprises an ordered, two-dimensional arrangement of at least 5 array elements, wherein each array element comprises at least one capture protein specific for a certain type of specified protein, and wherein at least two array elements are specific for different types of specified protein, wherein at least one type of specified protein in the solution binds to its respective capture protein;
b) adding a solution of one or more types of detection protein to the membrane, wherein at least one type of detection protein in the solution is capable of binding the bound specified protein; and
c) detecting bound detection protein, the detection of bound detection protein in a position on the microarray corresponding to an array element having a capture protein specific for a certain type of specified protein indicating the presence of that type of specified protein. - View Dependent Claims (73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95)
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Specification