%0 Journal Article %J PLoS One %D 2012 %T Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy %A Nath, Sangeeta %A Spencer, Virginia A %A Ju Han %A Hang Chang %A Zhang, Kai %A Fontenay, Gerald V %A Anderson, Charles %A Hyman, Joel M %A Nilsen-Hamilton, Marit %A Chang, Young-Tae %A Parvin, Bahram %K Animals %K Arabidopsis %K Cell Line %K Cell Survival %K Combinatorial Chemistry Techniques %K Fluorescent Dyes %K Humans %K Ligands %K Mice %K Microscopy %K Small Molecule Libraries %X

INTRODUCTION: Compounds exhibiting low non-specific intracellular binding or non-stickiness are concomitant with rapid clearing and in high demand for live-cell imaging assays because they allow for intracellular receptor localization with a high signal/noise ratio. The non-stickiness property is particularly important for imaging intracellular receptors due to the equilibria involved.

METHOD: Three mammalian cell lines with diverse genetic backgrounds were used to screen a combinatorial fluorescence library via high throughput live cell microscopy for potential ligands with high in- and out-flux properties. The binding properties of ligands identified from the first screen were subsequently validated on plant root hair. A correlative analysis was then performed between each ligand and its corresponding physiochemical and structural properties.

RESULTS: The non-stickiness property of each ligand was quantified as a function of the temporal uptake and retention on a cell-by-cell basis. Our data shows that (i) mammalian systems can serve as a pre-screening tool for complex plant species that are not amenable to high-throughput imaging; (ii) retention and spatial localization of chemical compounds vary within and between each cell line; and (iii) the structural similarities of compounds can infer their non-specific binding properties.

CONCLUSION: We have validated a protocol for identifying chemical compounds with non-specific binding properties that is testable across diverse species. Further analysis reveals an overlap between the non-stickiness property and the structural similarity of compounds. The net result is a more robust screening assay for identifying desirable ligands that can be used to monitor intracellular localization. Several new applications of the screening protocol and results are also presented.

%B PLoS One %V 7 %P e28802 %8 2012 %G eng %N 1 %R 10.1371/journal.pone.0028802