Performance Analysis of Non-Interferometry Based Surface Plasmon Resonance Microscopes

Surface plasmon microscopy has been of interest to the science and engineering community and has been utilized in broad aspects of applications and studies, including biochemical sensing and biomolecular binding kinetics. The benefits of surface plasmon microscopy include label-free detection, high sensitivity, and quantitative measurements. Here, a theoretical framework to analyze and compare several non-interferometric surface plasmon microscopes is proposed. The scope of the study is to (1) identify the strengths and weaknesses in each surface plasmon microscopes reported in the literature; (2) quantify their performance in terms of spatial imaging resolution, imaging contrast, sensitivity, and measurement accuracy for quantitative and non-quantitative imaging modes of the microscopes. Six types of non-interferometric microscopes were included in this study: annulus aperture scanning, half annulus aperture scanning, single-point scanning, double-point scanning, single-point scanning, at 45 degrees azimuthal angle, and double-point scanning at 45 degrees azimuthal angle. For non-quantitative imaging, there is a substantial tradeoff between the image contrast and the spatial resolution. For the quantitative imaging, the half annulus aperture provided the highest sensitivity of 127.058 rad/μm2 RIU−1, followed by the full annulus aperture of 126.318 rad/μm2 RIU−1. There is a clear tradeoff between spatial resolution and sensitivity. The annulus aperture and half annulus aperture had an optimal resolution, sensitivity, and crosstalk compared to the other non-interferometric surface plasmon resonance microscopes. The resolution depends strongly on the propagation length of the surface plasmons rather than the numerical aperture of the objective lens. For imaging and sensing purposes, the recommended microfluidic channel size and protein stamping size for surface plasmon resonance experiments is at least 25 μm for accurate plasmonic measurements.

Advanced Label-Free Laser Scanning Microscopy and Its Biological Imaging Application

By eliminating the photodamage and photobleaching induced by high intensity laser and fluorescent molecular, the label-free laser scanning microscopy shows powerful capability for imaging and dynamic tracing to biological tissues and cells. In this review, three types of label-free laser scanning microscopies: laser scanning coherent Raman scattering microscopy, second harmonic generation microscopy and scanning localized surface plasmon microscopy are discussed with their fundamentals, features and recent progress. The applications of label-free biological imaging of these laser scanning microscopies are also introduced. Finally, the performance of the microscopies is compared and the limitation and perspectives are summarized.