Kinase activity detection methods
OwnerLaurie L. Parker, Andrew M. Lipchikinventors
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Abstract
A strategy to take advantage of time-resolved luminescence of Ln3+-chelated phosphotyrosine-containing peptides, which facilitate efficient energy transfer to small molecule fluorophores conjugated to the peptides to produce orthogonally-colored biosensors for two different kinases is provided. The method enables multiplexed detection with high signal to noise in a high-throughput-compatible format and a platform that could be applied to other lanthanide metal and fluorophore combinations to achieve even greater multiplexing without the need for phosphospecific antibodies.
Claims17
A method for detecting the activities of two or more kinases comprising:
- a) contacting a first kinase and a second kinase with a first peptide and a second peptide, wherein:
- i) the first peptide is a substrate for the first kinase;
- ii) the second peptide is a substrate for the second kinase;
- iii) each peptide is associated with a lanthanide;
- iv) each peptide comprises a group capable of sensitizing the lanthanide that is associated with that peptide;
- and
- v) each peptide is linked to a fluorphore
- under conditions such that a first signal associated with the activity of the first kinase and a second signal that is associated with the activity of the second kinase are generated;
- and
- b) detecting the first signal and the second signal.
Dependencies are inferred from the claim text.