A surface plasmon enhanced FLIM-FRET imaging approach based on Au nanoparticles

Yinan Zhang,Yu Chen,Jun Yu,David Js Birch

doi:10.15761/mdde.1000119

Abstract

In this report we have demonstrated a fluorescence resonant energy transfer (FRET)-fluorescence lifetime imaging microscopy (FLIM) combined approach to study the intracellular pathway of gold nanoparticles. The detected energy transfer between gold nanorods (GNRs) and green fluorescence protein (GFP) labeled Hela cell earlyendosomes and the in-depth lifetime distribution analysis on the transfer process suggest an endocytotic uptake process of GNRs. Furthermore, the FRET-FLIM method profits from a surface plasmon enhanced energy transfer mechanism when taking into consideration of GNRs and two photon excitation, and is effective in biological imaging, sensing, and even in single molecular tracing in both in vivo and in vitro studies.

Highlights

Due to their unique physical and chemical properties, gold nanoparticles have been widely applied in in vivo and in vitro biological research, such as drug delivery [1-5], cancer therapy [6-13], imaging and sensing [14-20]
Multilayer coated nanoparticles were prepared following a layer by layer procedure: capped nanorods were centrifuged at 8500 rpm for 10 minutes, and were modified to optical density (O.D.) 1.0; nanorods solution was mixed with polystryrenesulfonate (PSS) solution (10mg/ ml in NaCl) with volume 5:1 and stirred for 5 minutes; after centrifuged at 8500 rpm for 15 minutes to remove excess polymers, and the remaining PSS coated nanorods solution was modified to O.D. 1.0; PSS coated particles were mixed with poly-diallyldimehtylammonium chloride (PDDAC) solution (10mg/ml in NaCl) with volume ratio 5:1 and stirred for another 5 minutes; excess polymers can be removed by centrifuge
In the two-photon excitation experiments, the decay curves can be fitted by a two-exponential model, in which the shorter component is from gold nanorods and the longer one from Fluor[405]

Summary

Introduction

Due to their unique physical and chemical properties, gold nanoparticles have been widely applied in in vivo and in vitro biological research, such as drug delivery [1-5], cancer therapy [6-13], imaging and sensing [14-20]. Understanding the intracellular pathways of gold nanoparticles and uptake mechanism is essential for all these applications, as these studies will provide information on uptake rate, route, final intracellular location, and the effects of nanoparticles to cell organelles, etc. This is not a straightforward task, because the properties of cells, such as cell groups and types, and characteristics of nanoparticles, such as the size, shape, surface charge and coating conditions, will all play a role effects on the uptake process [21-28]. While electron microscopes, such as transmission electron microscopy (TEM), can provide higher spacial resolution, but the specimen has to be treated at vacuum, is not possible to provide in situ, time lapse examination [40,41]

Methods

Results

Conclusion