title: Time-gated luminescence assay using nonmetal probes for determination of protein kinase activity-based disease markers.
abstract: A novel nonmetal optical probe ARC-1063 whose long-lifetime luminescence is induced by association with the target protein kinase is used for the measurement of the concentration of catalytic subunit of protein kinase A (PKAc) in complicated biological solutions. High affinity (K(D) = 10 pM toward PKAc) and unique optical properties of the probe enable its application for the measurement of picomolar concentrations of PKAc in the presence of high concentrations of other proteins. The described assay is applicable in the high-throughput format with the instrument setups designed for lanthanide-based time-gated (time-resolved) luminescence methods. The assay is used for demonstration that extracellular PKAc (ECPKA) is present in plasma samples of all healthy persons and cancer patients but great care must be taken for procedures of treatment of blood samples to avoid disruption, damage, or activation of platelets in the course of plasma (or serum) preparation and conservation.
authors.affiliation: Institute of Chemistry, University of Tartu, Ravila 14a, 50411 Tartu, Estonia.
authors.forename: Marje
authors.initials: M
authors.lastname: Kasari
authors.forename: Peeter
authors.initials: P
authors.lastname: Padrik
authors.forename: Angela
authors.initials: A
authors.lastname: Vaasa
authors.forename: Kristi
authors.initials: K
authors.lastname: Saar
authors.forename: Krista
authors.initials: K
authors.lastname: Leppik
authors.forename: Jaan
authors.initials: J
authors.lastname: Soplepmann
authors.forename: Asko
authors.initials: A
authors.lastname: Uri
mesh_terms: D015415:Biomarkers / Q000097:blood*; D001792:Blood Platelets / Q000166:cytology; D001943:Breast Neoplasms / Q000097:blood*; D016022:Case-Control Studies; D020134:Catalytic Domain; D015179:Colorectal Neoplasms / Q000097:blood*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000097:blood*; D004957:Estonia; D005260:Female; D057166:High-Throughput Screening Assays; D006801:Humans; D007700:Kinetics; D028581:Lanthanoid Series Elements; D057230:Limit of Detection; D049449:Luminescence; D008163:Luminescent Measurements / Q000379:methods*; D015539:Platelet Activation; D013048:Specimen Handling
chemical_list: D015415:Biomarkers; D028581:Lanthanoid Series Elements; D017868:Cyclic AMP-Dependent Protein Kinases