Methods and compositions for detecting the activation state of multiple proteins in single cells
AssigneeThe Board of Trustees of the Leland Stanford Junior University
InventorsGarry P. Nolan, Omar D. Perez
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Abstract
The invention provides methods and compositions for simultaneously detecting the activation state of a plurality of proteins in single cells using flow cytometry. The invention further provides methods and compositions of screening for bioactive agents capable of coordinately modulating the activity of a plurality of proteins in single cells. The methods and compositions can be used to determine the protein activation profile of a cell for predicting or diagnosing a disease state, and for monitoring treatment of a disease state.
Claims32
A method of detecting the activation state of single cells, said method comprising the steps of:
- a) providing a cell comprising a first and a second activatable protein, wherein said first and second activatable proteins are different proteins that each have at least an activated isoform, and a non-activated isoform;
- b) contacting said cell with at least two activation state-specific antibodies,
- wherein a first of said at least two activation state-specific antibodies is specific for said activated isoform of said first activatable protein;
- and a second of said at least two activation state-specific antibodies is specific for said activated isoform of said second activatable protein, wherein said two activation state-specific antibodies are distinguishably detectable;
- and
- c) detecting a presence or absence of binding of said first and said second activation state-specific antibodies to their corresponding activated isoform of said first and second activatable proteins in said cell, wherein said presence or absence of binding of said first activation state-specific antibody is indicative of the activation state of said first activatable protein, and said presence or absence of binding of said second activation state-specific antibody is indicative of the activation state of said second activatable protein.
A method of detecting the activation state of single cells, said method comprising the steps of:
- a) providing a cell comprising a first and a second activatable protein, wherein said first and second activatable proteins are different proteins that each have at least an activated isoform and a non-activated isoform;
- b) contacting said cell with at least two activation state-specific binding agents, wherein a first of said at least two activation state-specific binding agents is specific for said activated isoform of said first activatable protein and a second of said at least two activation state-specific binding agents is specific for said activated isoform of said second activatable protein, wherein said two activation state-specific binding agents are distinguishably detectable;
- and
- c) detecting a presence or absence of binding of said first and said second activation state-specific binding agents to their corresponding activated isoform of said first and second activatable proteins in said single cell, wherein said presence or absence of binding of said first activation state-specific binding agent is indicative of the activation state of said first activatable protein, and said presence or absence of binding of said second activation state-specific binding agent is indicative of the activation state of said second activatable protein.
Dependencies are inferred from the claim text.