Simultaneous detection of intracellular target and off-target binding of small molecule cancer drugs at nanomolar concentrations.

ABSTRACT:

Background and purpose

In vitro assays that determine activities of drug candidates with isolated targets have only limited predictive value for activities in cellular assays. Poor membrane permeability and off-target binding are major reasons for such discrepancies. However, it still difficult to directly analyse off-target binding at the same time as target binding, on a subcellular level. Here, we present a combination of fluorescence correlation spectroscopy (FCS) and fluorescence cross-correlation spectroscopy (FCCS) as a solution to this problem.

Experimental approach

The well-established dihydrofolate reductase inhibitor methotrexate and the kinase inhibitors PD173956 and purvalanol B were conjugated via polyethylene glycol linkers with the fluorophore Cy5. The cellul

SUBMITTER: Glauner H 

PROVIDER: S-EPMC2936001 | biostudies-literature | 2010 Jun

REPOSITORIES: biostudies-literature