Visualization of lipids and proteins at high spatial and temporal resolution via interferometric scattering (iSCAT) microscopy

Susann Spindler,Isabel D Alves,Sébastien Lecommandoux,Tristan Nowak,Elisabeth Garanger,Katharina König,Vahid Sandoghdar,Marek Piliarik,Hannah E Stein,Jens Ehrig,Richard W Taylor

doi:10.1088/0022-3727/49/27/274002

Abstract

Microscopy based on the interferometric detection of light scattered from nanoparticles (iSCAT) was introduced in our laboratory more than a decade ago. In this work, we present various capabilities of iSCAT for biological studies by discussing a selection of our recent results. In particular, we show tracking of lipid molecules in supported lipid bilayers (SLBs), tracking of gold nanoparticles with diameters as small as 5 nm and at frame rates close to 1 MHz, 3D tracking of Tat peptide-coated nanoparticles on giant unilamellar vesicles (GUVs), imaging the formation of lipid bilayers, sensing single unlabelled proteins and tracking their motion under electric fields, as well as challenges of studying live cell membranes. These studies set the ground for future quantitative research on dynamic biophysical processes at the nanometer scale.

Highlights

The arsenal of light microscopy consists of a number of different contrast mechanisms such as bright field, dark field, and phase contrast, optical approaches
We recently demonstrated tracking of GM1 lipids within supported lipid bilayers (SLBs) of DOPC on glass at a spatiotemporal resolution superior to that achieved with fluorescence [1]
In the lower inset of figure 3(a) we show a background-corrected interferometric detection of light scattered from nanoparticles (iSCAT) image of a 5 nm gold nanoparticle (GNP) bound via streptavidin to headgroup-biotinylated DOPE in a DOPC SLB

Summary

Introduction

A fundamental understanding of structure and dynamics in biological systems requires measurement methods that offer. The precision of localizing the particle is dependent on the achievable signal-to-noise ratio (SNR) and the number of detected photons [1, 2] This in turn demands a certain length of integration time. The strength of the reference beam determines the contrast, the sensitivity of the method and the attainable shot-noise-limited SNR depends fundamentally on α [5]. The latter is proportional to the quantity (εp − εm)/(εp + 2εm) with εp and εm denoting the dielectric functions of the particle and its surrounding medium, respectively. It turns out that this method allows one to detect even a single small unlabelled protein [10]

Experimental setup

Tracking 2D lipid diffusion on a supported lipid bilayer

Visualizing lipid bilayer formation from single lipid vesicles