Interferometric Scattering (iSCAT) Microscopy and Related Techniques

Abstract

Interferometric scattering (iSCAT) microscopy is a powerful tool for label-free sensitive detection and imaging of nanoparticles to high spatio-temporal resolution. As it was born out of detection principles central to conventional microscopy, we begin by surveying the historical development of the microscope to examine how the exciting possibility for interferometric scattering microscopy with sensitivities sufficient to observe single molecules has become a reality. We discuss the theory of interferometric detection and also issues relevant to achieving a high detection sensitivity and speed. A showcase of numerous applications and avenues of novel research across various disciplines that iSCAT microscopy has opened up is also presented.1

Interferometric Scattering Microscopy: Seeing Single Nanoparticles and Molecules via Rayleigh Scattering

“…1 This led many scientists to invent clever strategies such as dark-field, phase contrast, and differential interference contrast (DIC) microscopies. 1 Interestingly, however, these methods did not push the limits of detection sensitivity with regard to the achievable signal-to-noise ratio (SNR). Instead, they merely provided convenient ways of identifying image features within the limitations of the available detectors.…”

“…Some of the technical difficulties in doing so involve eliminating the temporal fluctuations in the illumination and addressing the limited dynamic range of the detector. 1…”

“…In this Mini Review, we discuss the developments of the past two decades, which have led to the label-free detection of nanoparticles including single dye molecules, semiconductor quantum dots, viruses, and small proteins through interferometric scattering (iSCAT) microscopy. A thorough account can be found in ref (1).…”

Measuring single‐cell protein secretion in immunology: Technologies, advances, and applications

“…Unmatched in sensitivity (single‐protein resolution), iSCAT microscopy has great potential but is also a long way from widespread use. Its primary disadvantage remains its throughput since only one individual cell can be analyzed at a time [25].…”

TRAIT2D: a Software for Quantitative Analysis of Single Particle Diffusion Data

“…Among the earliest example of SPT experiments, we can mention the testing of Einstein's theory of Brownian motion (2). The introduction of electronic camera detection and innovative microscopy systems have made it possible to collect SPT data with framerates up to 50kHz (3)(4)(5)(6). While seemingly straightforward, the analysis of SPT data is not immune from potential pitfalls.…”