Advanced Search

Journal Navigation

Journal Home

Subscriptions

Archive

Contact Us

Table of Contents

CiteULike is a free service for managing and discovering scholarly references - click here to get started.

Sign In to gain access to subscriptions and/or personal tools.
Journal of Biomolecular Screening
This Article
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
1087057106286402v1
11/4/413    most recent
Right arrow References
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Saved Citations
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Request Reprints
Right arrow Add to My Marked Citations
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Right arrow Citing Articles via Scopus
Google Scholar
Right arrow Articles by Stankewicz, C.
Right arrow Articles by Rininsland, F. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Stankewicz, C.
Right arrow Articles by Rininsland, F. H.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

A Robust Screen for Inhibitors and Enhancers of Phosphoinositide-3 Kinase (PI3K) Activities by Ratiometric Fluorescence Superquenching

Casey Stankewicz

QTL Biosystems, Santa Fe, NM

Frauke H. Rininsland

QTL Biosystems, Santa Fe, NM

Aberrant regulation of phosphoinositide 3-kinase (PI3K) activity is implicated in various diseases such as cancer and diabetes. Thus, high-throughput screening (HTS) of small-molecule inhibitors for PI3 kinases is an appealing strategy for drug development. Despite the attractiveness of lipid kinases as drug targets, screening for inhibitors for PI3K activities has been hampered by limited assay formats adaptable for HTS. The authors describe a homogeneous, direct, and nonradioactive assay for highly sensitive detection of PI3K{alpha}, ß, {delta}, and {gamma} activities, which is suitable for HTS. The assay is based on fluorescence superquenching of a conjugated polymer upon metal-ion-mediated association of phosphorylated and dye-labeled substrates. As a result of phosphorylation, quencher and polymer are brought into proximity, and fluorescent energy transfer occurs. This event can be monitored as either fluorescence quench of the polymer or as enhanced emission from the quencher. Ratiometric analysis of the wavelengths eliminates interferences from autofluorescing compounds, which are present in HTS libraries. The platform has been adapted for the 384-well microplate format and delivers Z factors of > 0.6 at substrate conversions as low as 7%. Using this assay platform, several unreported inhibitors and activators of PI3Ks were identified in an 84- compound screen.

Key Words: cancer • diabetes • phosphoinositide-3 kinases • high-throughput screening • superquenching

This version was published on June 1, 2006

Journal of Biomolecular Screening, Vol. 11, No. 4, 413-422 (2006)
DOI: 10.1177/1087057106286402
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?


This article has been cited by other articles:


Home page
 J Biomol ScreenHome page
B. Boldyreff, T. L. Rasmussen, H. H. Jensen, A. Cloutier, L. Beaudet, P. Roby, and O.-G. Issinger
Expression and Purification of PI3 Kinase {alpha} and Development of an ATP Depletion and an AlphaScreen PI3 Kinase Activity Assay
J Biomol Screen, December 1, 2008; 13(10): 1035 - 1040.
[Abstract] [PDF]

Home page
 J Biomol ScreenHome page
T. A. Klink, K. M. Kleman-Leyer, A. Kopp, T. A. Westermeyer, and R. G. Lowery
Evaluating PI3 Kinase Isoforms Using TranscreenerTM ADP Assays
J Biomol Screen, July 1, 2008; 13(6): 476 - 485.
[Abstract] [PDF]