Development of a Transcreener kinase assay for protein kinase A and demonstration of concordance of data with a filter-binding assay format.
A Transcreener kinase fluorescence polarization (FP) assay has been developed for the serine/threonine kinase protein kinase A (PKA). The PKA Transcreener kinase assay is an homogenous, competitive antibody-based FP assay that uses Far Red Alexa Fluor 633-labeled adenosine 5' disphosphate (ADP) tracer and mouse monoclonal anti-ADP antibody. The Transcreener PKA assay was validated with both known PKA inhibitors and library compounds. The Transcreener PKA assay is resistant to low-wavelength (or common) fluorescent interference from small-molecule library compounds and generates IC50 results comparable with current radioactive filter-binding assay.
Huss
Karen L
KL
Lilly Research Laboratories A Division of Eli Lilly and Company, Indianapolis, Indiana 46285, USA. huss_karen_l@lilly.com
Blonigen
Pauline E
PE
Campbell
Robert M
RM
D000911:Antibodies, Monoclonal / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D016231:Fluorescence Polarization Immunoassay / Q000295:instrumentation / Q000379:methods; D011485:Protein Binding
D000911:Antibodies, Monoclonal; D017868:Cyclic AMP-Dependent Protein Kinases