Investigating Spatial Heterogeneity of Nanoparticles Movement in Live Cells with Pair-Correlation Microscopy and Phasor Analysis.

How nanoparticles distribute in living cells and overcome cellular barriers are important criteria in the design of drug carriers. Pair-correlation microscopy is a correlation analysis of fluctuation in the fluorescence intensity obtained by a confocal line scan that can quantify the dynamic properties of nanoparticle diffusion including the number of mobile nanoparticles, diffusion coefficient, and transit time across a spatial distance. Due to the potential heterogeneities in nanoparticle properties and the complexity within the cellular environment, quantification of averaged auto- and pair-correlation profiles may obscure important insights into the ability of nanoparticles to deliver drugs. To overcome this issue, we used phasor analysis to develop a data standardizing method, which can segment the scanned line into several subregions according to diffusion and address the spatial heterogeneity of nanoparticles moving inside cells. The phasor analysis is a fit-free method that represents autocorrelation profiles for each pixel relative to free diffusion on the so-called phasor plots. Phasor plots can then be used to select subpopulations for which the auto- and pair-correlation analysis can be performed separately. We demonstrate the phasor analysis for pair-correlation microscopy for investigating 16 nm, Cy5-labeled silica nanoparticles diffusing across the plasma membrane and green fluorescent proteins (GFP) diffusing across nuclear envelope in MCF-7 cells.

The use of a personal glucose meter for detecting procalcitonin through glucose encapsulated within liposomes.

Herein, a glucose meter-based immunosensing platform is developed that allows the quantification of procalcitonin (PCT) in whole blood samples. PCT is a biomarker for sepsis and its early detection would improve the safety of the patient, as the diagnostic process will be easier and faster. The method employs liposomes with encapsulated glucose as a signal generation tag, which are then used in a sandwich immunoassay by conjugating an antibody to the liposome. The optimal liposomes' size and concentration of encapsulated glucose is determined experimentally to be 200 nm and 27.8 mM, respectively. Upon the addition of a surfactant (Triton X-100), the glucose is released and a signal is detected with a personal glucose meter (PGM). This signal is directly proportional to the concentration of the PCT in the sample. The dynamic range of the assay developed was 0.153-15.38 nM, and could allow the detection of PCT as low as 0.15 nM. The assay showed a high selectivity toward PCT against other proteins such as C-reactive protein and human serum albumin and good reproducibility. This assay was able to quantitatively determine the amount of PCT in whole blood samples at clinically-relevant concentrations.