Multiparametric Biosensors for Characterizing Extracellular Vesicle Subpopulations.

Extracellular vesicles (EVs) are an important intercellular communication conduit for cells that have applications in precision therapy and targeted drug delivery. Small EVs, or exosomes, are a 30-150 nm phospholipid-encased subpopulation of EVs that are particularly difficult to characterize due to their small size and because they are difficult to isolate using conventional methods. In this review, we discuss some recent advances in exosome isolation, purification, and sensing platforms using microfluidics, acoustics, and size exclusion chromatography. We discuss some of the challenges and unanswered questions with respect to understanding exosome size heterogeneity and how modern biosensor technology can be applied to exosome isolation. In addition, we discuss how some advancements in sensing platforms such as colorimetric, fluorescent, electronic, surface plasmon resonance (SPR), and Raman spectroscopy may be applied to exosome detection in multiparametric systems. The application of cryogenic electron tomography and microscopy to understanding exosome ultrastructure will become vital as this field progresses. In conclusion, we speculate on some future needs in the exosome research field and how these technologies could be applied.

A Molecularly Imprinted Sol-Gel Electrochemical Sensor for Naloxone Determination.

A molecularly imprinted sol-gel is reported for selective and sensitive electrochemical determination of the drug naloxone (NLX). The sensor was developed by combining molecular imprinting and sol-gel techniques and electrochemically grafting the sol solution onto a functionalized multiwall carbon nanotube modified indium-tin oxide (ITO) electrode. The sol-gel layer was obtained from acid catalyzed hydrolysis and condensation of a solution composed of triethoxyphenylsilane (TEPS) and tetraethoxysilane (TES). The fabrication, structure and properties of the sensing material were characterized via scanning electron microscopy, spectroscopy and electrochemical techniques. Parameters affecting the sensor's performance were evaluated and optimized. A sensor fabricated under the optimized conditions responded linearly between 0.0 µM and 12 µM NLX, with a detection limit of 0.02 µM. The sensor also showed good run-to-run repeatability and batch-to-batch performance reproducibility with relative standard deviations (RSD) of 2.5-7.8% (n = 3) and 9.2% (n = 4), respectively. The developed sensor displayed excellent selectivity towards NLX compared to structurally similar compounds (codeine, fentanyl, naltrexone and noroxymorphone), and was successfully used to measure NLX in synthetic urine samples yielding recoveries greater than 88%.

Size-based proteins separation using polymer-entrapped colloidal self-assembled nanoparticles on-chip.

We report on a facile method to stabilize colloidal self-assembled (CSA) nanoparticles packed in microchannels for high speed size-based separation of proteins. Silica nanoparticles, self-assembled in a network of microfluidic channels, were stabilized with a methacrylate polymer prepared in situ through photopolymerization. The entrapment conditions were investigated to minimize the effect of the polymer matrix on the structure of the packing and the separation properties of the CSA beds. SEM shows that the methacrylate matrix links the nanoparticles at specific sphere-sphere contact points, improving the stability of the CSA structure at high electric fields (up to at least 1800 V/cm), allowing fast and efficient separation. The %RSD of the protein migration times varied between 0.3 and 0.5% (n = 4, in 1 day) and <0.83% over a period of 7 days (n = 28 runs) in a single device, at high field strength. The overall %RSD of protein migration times from chip-to-chip across a single fabrication run was 4.3% (n = 3) and between fabrication runs was 11% (n = 35), with 87% fabrication yield, demonstrating reproducible packing and entrapment behavior. The optimized entrapped CSA beds demonstrated better separation performance (plate height, H ∼ 200 nm) than similarly prepared on-chip CSA beds without the polymer entrapment. Polymer-entrapped CSA beds also exhibited superior protein resolving power: the minimum resolvable molecular weight difference of proteins in the polymer-entrapped CSA bed is 0.6 kDa versus ∼9 kDa for the native silica CSA bed (i.e. without polymer entrapment).

Novel molecularly imprinted polymers for the selective extraction and determination of metoclopramide in human serum and urine samples using high-performance liquid chromatography.

This work was performed in order to study the possibilities in using molecularly imprinted polymers (MIPs) as sorbent material in solid-phase extraction (MISPE) for the sample clean-up technique for the determination of metoclopramide (MCP) in biological fluids. The effective factors influencing the bulk polymerization have been studied. Molecular recognition properties, binding capability and selectivity of the molecularly imprinted polymers (MIPs) were evaluated and the results revealed the obtained MIPs have high affinity for MCP in aqueous environment. The optimal conditions for solid-phase extraction (SPE) consisted of conditioning with 1 mL of methanol and 1 mL of deionized water at neutral pH, loading with 1 mL of the water sample (50 microg L(-1)) at pH 8.5, washing using 1 mL of acetone and elution with 2x 1 mL methanol/acetic acid (10/1, v/v). After optimization of SPE procedure, the MIP was then directly used to selectively extract the target drug from human serum and urine with an extraction recovery of more than 90%. Chromatograms of the eluate solutions show an efficient clean-up, which supports the potential of MISPE for clean-up of trace amounts of MCP from serum and urine. The limits of detection of MCP in human serum and urine were 3 and 1.2 microg L(-1), respectively.