Surface plasmon-assisted microscope

Julian Borejdo, Zygmunt Gryczynski, Rafal Fudala, Chaitanya R. Joshi, Kathleen Borgmann, Anuja Ghorpade, Ignacy Gryczynski

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Total internal reflection microscopy (TIRF) has been a powerful tool in biological research. The most valuable feature of the method has been the ability to image 100-to 200-nm-thick layer of cell features adjacent to a coverslip, such as membrane lipids, membrane receptors, and structures proximal-to-basal membranes. Here, we demonstrate an alternative method of imaging thin-layer proximal-to-basal membranes by placing a sample on a high refractive index coverslip covered by a thin layer of gold. The sample is illuminated using the Kretschmann method (i.e., from the top to an aqueous medium). Fluorophores that are close to the metal surface induce surface plasmons in the metal film. Fluorescence from fluorophores near the metal surface couple with surface plasmons allowing them to penetrate the metal surface and emerge at a surface plasmon coupled emission angle. The thickness of the detection layer is further reduced in comparison with TIRF by metal quenching of fluorophores at a close proximity (below 10 nm) to a surface. Fluorescence is collected by a high NA objective and imaged by EMCCD or converted to a signal by avalanche photodiode fed by a single-mode optical fiber inserted in the conjugate image plane of the objective. The system avoids complications of through-the-objective TIRF associated with shared excitation and emission light path, has thin collection thickness, produces excellent background rejection, and is an effective method to study molecular motion.

Original languageEnglish
Article number060502
JournalJournal of Biomedical Optics
Volume23
Issue number6
DOIs
StatePublished - 1 Jun 2018

Fingerprint

Microscopes
microscopes
membranes
metal surfaces
Metals
microscopy
Fluorophores
plasmons
Microscopic examination
Plasmons
fluorescence
Membranes
metal films
rejection
avalanches
photodiodes
light emission
lipids
proximity
Fluorescence

Keywords

  • fluorescence
  • microscopy
  • surface plasmon coupled emission

Cite this

Borejdo, Julian ; Gryczynski, Zygmunt ; Fudala, Rafal ; Joshi, Chaitanya R. ; Borgmann, Kathleen ; Ghorpade, Anuja ; Gryczynski, Ignacy. / Surface plasmon-assisted microscope. In: Journal of Biomedical Optics. 2018 ; Vol. 23, No. 6.
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Surface plasmon-assisted microscope. / Borejdo, Julian; Gryczynski, Zygmunt; Fudala, Rafal; Joshi, Chaitanya R.; Borgmann, Kathleen; Ghorpade, Anuja; Gryczynski, Ignacy.

In: Journal of Biomedical Optics, Vol. 23, No. 6, 060502, 01.06.2018.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Gryczynski, Zygmunt

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