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Time-resolved surface-enhanced ellipsometric contrast imaging for label-free analysis of biomolecular recognition reactions on glycolipid domains

Gunnarsson, Anders LU ; Bally, Marta; Jönsson, Peter LU ; Médard, Nicolas and Höök, Fredrik LU (2012) In Analytical Chemistry 84(15). p.45-6538
Abstract

We have applied surface-enhanced ellipsometry contrast (SEEC) imaging for time-resolved label-free visualization of biomolecular recognition events on spatially heterogeneous supported lipid bilayers (SLB). Using a conventional inverted microscope equipped with total internal reflection (TIR) illumination, biomolecular binding events were monitored with a lateral resolution near the optical diffraction limit at an acquisition rate of ~1 Hz with a sensitivity in terms of surface coverage of ~1 ng/cm(2). Despite the significant improvement in spatial resolution compared to alternative label-free surface-based imaging technologies, the sensitivity remains competitive with surface plasmon resonance (SPR) imaging and imaging ellipsometry.... (More)

We have applied surface-enhanced ellipsometry contrast (SEEC) imaging for time-resolved label-free visualization of biomolecular recognition events on spatially heterogeneous supported lipid bilayers (SLB). Using a conventional inverted microscope equipped with total internal reflection (TIR) illumination, biomolecular binding events were monitored with a lateral resolution near the optical diffraction limit at an acquisition rate of ~1 Hz with a sensitivity in terms of surface coverage of ~1 ng/cm(2). Despite the significant improvement in spatial resolution compared to alternative label-free surface-based imaging technologies, the sensitivity remains competitive with surface plasmon resonance (SPR) imaging and imaging ellipsometry. The potential of the technique to discriminate local differences in protein binding kinetics was demonstrated by time-resolved imaging of anti-GalCer antibodies binding to phase-separated lipid bilayers consisting of phosphatidylcholine (POPC) and galactosylceramide (GalCer). A higher antibody binding capacity was observed on the GalCer-diluted fluid region in comparison to the GalCer-rich gel phase domains. This observation is tentatively attributed to differences in the presentation of the GalCer epitope in the two phases, resulting in differences in availability of the ligand for antibody binding. The complementary information obtained by swiftly switching between SEEC and fluorescence (including TIR fluorescence) imaging modes was used to support the data interpretation. The simplicity and generic applicability of the concept is discussed in terms of microfluidic applications.

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author
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Antibodies, Fluorescence Recovery After Photobleaching, Galactosylceramides, Kinetics, Lipid Bilayers, Microfluidic Analytical Techniques, Microscopy, Phosphatidylcholines, Protein Binding, Surface Properties, Time Factors, Journal Article
in
Analytical Chemistry
volume
84
issue
15
pages
8 pages
publisher
The American Chemical Society
external identifiers
  • scopus:84864577740
ISSN
1520-6882
DOI
10.1021/ac300832k
language
English
LU publication?
no
id
ab1a16a8-607f-4d61-bcb7-80a51be4c9cb
date added to LUP
2018-01-26 10:29:51
date last changed
2018-07-08 04:29:46
@article{ab1a16a8-607f-4d61-bcb7-80a51be4c9cb,
  abstract     = {<p>We have applied surface-enhanced ellipsometry contrast (SEEC) imaging for time-resolved label-free visualization of biomolecular recognition events on spatially heterogeneous supported lipid bilayers (SLB). Using a conventional inverted microscope equipped with total internal reflection (TIR) illumination, biomolecular binding events were monitored with a lateral resolution near the optical diffraction limit at an acquisition rate of ~1 Hz with a sensitivity in terms of surface coverage of ~1 ng/cm(2). Despite the significant improvement in spatial resolution compared to alternative label-free surface-based imaging technologies, the sensitivity remains competitive with surface plasmon resonance (SPR) imaging and imaging ellipsometry. The potential of the technique to discriminate local differences in protein binding kinetics was demonstrated by time-resolved imaging of anti-GalCer antibodies binding to phase-separated lipid bilayers consisting of phosphatidylcholine (POPC) and galactosylceramide (GalCer). A higher antibody binding capacity was observed on the GalCer-diluted fluid region in comparison to the GalCer-rich gel phase domains. This observation is tentatively attributed to differences in the presentation of the GalCer epitope in the two phases, resulting in differences in availability of the ligand for antibody binding. The complementary information obtained by swiftly switching between SEEC and fluorescence (including TIR fluorescence) imaging modes was used to support the data interpretation. The simplicity and generic applicability of the concept is discussed in terms of microfluidic applications.</p>},
  author       = {Gunnarsson, Anders and Bally, Marta and Jönsson, Peter and Médard, Nicolas and Höök, Fredrik},
  issn         = {1520-6882},
  keyword      = {Antibodies,Fluorescence Recovery After Photobleaching,Galactosylceramides,Kinetics,Lipid Bilayers,Microfluidic Analytical Techniques,Microscopy,Phosphatidylcholines,Protein Binding,Surface Properties,Time Factors,Journal Article},
  language     = {eng},
  month        = {08},
  number       = {15},
  pages        = {45--6538},
  publisher    = {The American Chemical Society},
  series       = {Analytical Chemistry},
  title        = {Time-resolved surface-enhanced ellipsometric contrast imaging for label-free analysis of biomolecular recognition reactions on glycolipid domains},
  url          = {http://dx.doi.org/10.1021/ac300832k},
  volume       = {84},
  year         = {2012},
}