Postrecording Pixel-Reconstruction Approach for Correcting the Lateral Drifts in Surface Plasmon Resonance Microscope.

Surface plasmon resonance microscope (SPRM) sample stage inevitably suffers from lateral drifts as a result of many environmental factors including thermal fluctuation, mechanical vibration, and relaxation. It places great obstacles to time-lapsed imaging and measurements that need high spatial resolution or long recording time. Existing solutions often require experimental efforts such as the addition of optical markers together with piezoelectric stage-based active feedback configurations. Herein, we propose an all-digital, postrecording image-processing method to remove the lateral drift in a series of time-lapsed SPRM images. The method first calculates the value of lateral drift at subpixel accuracy by combining image cross-correlation analysis and superlocalization strategy. It subsequently reconstructed the drift-free image sequences in a pixel-by-pixel and frame-by-frame manner, according to the linear decomposition and reconstruction principle. This method purely relies on image processing, and it does not require any experimental efforts or hardware. In addition to SPRM, we further demonstrated the applicability of the present method in other types of optical imaging techniques including bright-field transmission microscope and dark-field scattering microscope.

Influence of Fixation and Permeabilization on the Mass Density of Single Cells: A Surface Plasmon Resonance Imaging Study.

Fixation and permeabilization of cells and tissues are essential processes in biological techniques like immunofluorescence and immunohistochemistry for cell biology studies. In typical procedures, the biological samples are treated by paraformaldehyde and Triton X-100 to achieve cellular fixation and permeabilization, respectively, prior to the incubation with specific antibodies. While it is well-known that the integrity of cell membrane has been broken during these processes, quantitative studies on the loss of cellular mass density and the enhancement of molecular accessibility at single cell level are still rare. In this study, we employed the surface plasmon resonance (SPR) imaging technique to monitor the mass density change of single cells during sequential fixation and permeabilization processes. We further utilize the osmotic responses of single cells to sugar molecules as an indicator to evaluate the integrity of cell membranes. It was found that, while fixation initially destructed the integrity of cell membranes and increased the permeability of intra- and extra-cellular molecules, it was permeabilization process that substantially induced significant loss in cellular mass density.

Optical Imaging of Phase Transition and Li-Ion Diffusion Kinetics of Single LiCoO(2) Nanoparticles During Electrochemical Cycling.

Understanding the phase transition and Li-ion diffusion kinetics of Li-ion storage nanomaterials holds promising keys to further improve the cycle life and charge rate of the Li-ion battery. Traditional electrochemical studies were often based on a bulk electrode consisting of billions of electroactive nanoparticles, which washed out the intrinsic heterogeneity among individuals. Here, we employ optical microscopy, termed surface plasmon resonance microscopy (SPRM), to image electrochemical current of single LiCoO2 nanoparticles down to 50 fA during electrochemical cycling, from which the phase transition and Li-ion diffusion kinetics can be quantitatively resolved in a single nanoparticle, in operando and high throughput manner. SPRM maps the refractive index (RI) of single LiCoO2 nanoparticles, which significantly decreases with the gradual extraction of Li-ions, enabling the optical read-out of single nanoparticle electrochemistry. Further scanning electron microscopy characterization of the same batch of nanoparticles led to a bottom-up strategy for studying the structure-activity relationship. As RI is an intrinsic property of any material, the present approach is anticipated to be applicable for versatile kinds of anode and cathode materials, and to facilitate the rational design and optimization toward durable and fast-charging electrode materials.

Determining the Absolute Concentration of Nanoparticles without Calibration Factor by Visualizing the Dynamic Processes of Interfacial Adsorption.

Previous approaches of determining the molar concentration of nanoparticles often relied on the calibration factors extracted from standard samples or required prior knowledge regarding the geometry, optical, or chemical properties. In the present work, we proposed an absolute quantification method that determined the molar concentration of nano-objects without any calibration factor or prior knowledge. It was realized by monitoring the dynamic adsorption processes of individual nanoparticles with a high-speed surface plasmon resonance microscopy. In this case, diffusing nano-objects stochastically collided onto an adsorption interface and stayed there ("hit-n-stay" scenario), resulting in a semi-infinite diffusion system. The dynamic processes were analyzed with a theoretical model consisting of Fick's laws of diffusion and random-walk assumption. The quantification of molar concentration was achieved on the basis of an analytical expression, which involved only physical constants and experimental parameters. By using spherical polystyrene nanoparticles as a model, the present approach provided a molar concentration with excellent accuracy.

Digitizing Gold Nanoparticle-Based Colorimetric Assay by Imaging and Counting Single Nanoparticles.

Gold colloid changes its color when the internanoparticle distance changes. On the basis of analyte-induced aggregation or disaggregation behavior of gold nanoparticles (AuNPs), versatile colorimetric assays have been developed for measuring various kinds of analytes including proteins, DNA, small molecules, and ions. Traditional read-out signals, which are usually measured by a spectrometer or naked eyes, are based on the averaged extinction properties of a bulk solution containing billions of nanoparticles. Averaged extinction property of a large amount of nanoparticles diminished the contribution from rare events when the analyte concentration was low, thus resulting in limited detection sensitivity. Instead of measuring the averaged optical property from bulk colloid, in the present work, we proposed a digital counterpart of the colorimetric assay by imaging and counting individual AuNPs. This method quantified the analyte concentration with the number percentage of large-sized AuNPs aggregates, which were digitally counted with surface plasmon resonance microscopy (SPRM), a plasmonic imaging technique recently developed by us and other groups. SPRM was able to identify rare AuNPs aggregates despite their small population and greatly improved the detection sensitivity as demonstrated by two model systems based on analyte-induced aggregation and disaggregation, respectively. Furthermore, besides plasmonic AuNPs, SPRM is also suitable for imaging and counting nonplasmonic nanomaterials such as silica and metal oxide with poor extinction properties. It is thus anticipated that the present digitized assay holds a great potential for expanding the colorimetric assay to broad categories of nonplasmonic nanoparticles.

Plasmonic imaging of electrochemical oxidation of single nanoparticles.

Measuring electrochemical activities of nanomaterials is critical for creating novel catalysts, for developing ultrasensitive sensors, and for understanding fundamental nanoelectrochemistry. However, traditional electrochemical methods measure a large number of nanoparticles, which wash out the properties of individual nanoparticles. We report here a study of transient electrochemical oxidation of single Ag nanoparticles during collision with an electrode and voltammetry of single nanoparticles immobilized on the electrode using a plasmonic-based electrochemical current microscopy. This technique images both electrochemical reaction and size of the same individual nanoparticle, enabling quantitative examination of size-dependent electrochemical activities at single nanoparticle level. The imaging capability further allows detection of the reaction kinetics of each individual nanoparticle and analysis of the average behaviors of multiple nanoparticles. The average kinetics and size dependence can be accurately described by the Tafel equation, but there is a large variability between different nanoparticles, which underscores the importance of single nanoparticle analysis.