Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy

Sangeeta Nath, Virginia A. Spencer, Ju Han, Hang Chang, Kai Zhang, Gerald V. Fontenay, Charles T. Anderson, Joel M. Hyman, Marit Nilsen-Hamilton, Young Tae Chang, Bahram Parvin

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

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.

Original languageEnglish (US)
Article numbere28802
JournalPloS one
Volume7
Issue number1
DOIs
StatePublished - Jan 5 2012

Fingerprint

binding properties
Microscopy
microscopy
Microscopic examination
Throughput
Ligands
Screening
Chemical compounds
chemical compounds
image analysis
screening
Imaging techniques
cells
Assays
Cells
cell lines
Fluxes
Cell Line
receptors
Plant Roots

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Nath, S., Spencer, V. A., Han, J., Chang, H., Zhang, K., Fontenay, G. V., ... Parvin, B. (2012). Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy. PloS one, 7(1), [e28802]. https://doi.org/10.1371/journal.pone.0028802
Nath, Sangeeta ; Spencer, Virginia A. ; Han, Ju ; Chang, Hang ; Zhang, Kai ; Fontenay, Gerald V. ; Anderson, Charles T. ; Hyman, Joel M. ; Nilsen-Hamilton, Marit ; Chang, Young Tae ; Parvin, Bahram. / Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy. In: PloS one. 2012 ; Vol. 7, No. 1.
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abstract = "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.",
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Nath, S, Spencer, VA, Han, J, Chang, H, Zhang, K, Fontenay, GV, Anderson, CT, Hyman, JM, Nilsen-Hamilton, M, Chang, YT & Parvin, B 2012, 'Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy', PloS one, vol. 7, no. 1, e28802. https://doi.org/10.1371/journal.pone.0028802

Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy. / Nath, Sangeeta; Spencer, Virginia A.; Han, Ju; Chang, Hang; Zhang, Kai; Fontenay, Gerald V.; Anderson, Charles T.; Hyman, Joel M.; Nilsen-Hamilton, Marit; Chang, Young Tae; Parvin, Bahram.

In: PloS one, Vol. 7, No. 1, e28802, 05.01.2012.

Research output: Contribution to journalArticle

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T1 - Identification of fluorescent compounds with non-specific binding property via high throughput live cell microscopy

AU - Nath, Sangeeta

AU - Spencer, Virginia A.

AU - Han, Ju

AU - Chang, Hang

AU - Zhang, Kai

AU - Fontenay, Gerald V.

AU - Anderson, Charles T.

AU - Hyman, Joel M.

AU - Nilsen-Hamilton, Marit

AU - Chang, Young Tae

AU - Parvin, Bahram

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N2 - 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.

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