Label-free SERS assay combined with multivariate spectral data analysis for lamotrigine quantification in human serum.

Considering the need for a more time and cost-effective method for lamotrigine (LTG) detection in clinics we developed a fast and robust label-free assay based on surface-enhanced Raman scattering (SERS) for LTG quantification from human serum. The optimization and application of the developed assay is presented  showing the: (i) exploration of different methods for LTG separation from human serum; (ii) implementation of a molecular adsorption step on an ordered Au nanopillar SERS substrate; (iii) adaptation of a fast scanning of the SERS substrate, performed with a custom-built compact Raman spectrometer; and (iv) development of LTG quantification methods with univariate and multivariate spectral data analysis. Our results showed, for the first time, the SERS-based characterization of LTG and its label-free identification in human serum. We found that combining a miniaturized solid phase extraction, as sample pre-treatment with the SERS assay, and using a multivariate model is an optimal strategy for LTG quantification in human serum in a linear range from 9.5 to 75 µM, with LoD and LoQ of 3.2 µM and 9.5 µM, respectively, covering the suggested clinical therapeutic window. We also showed that the developed assay allowed for quantifying LTG from human serum in the presence of other drugs, thereby demonstrating the robustness of label-free SERS. The sensing approach and instrumentation can be further automated and integrated in devices that can advance the drug monitoring in real clinical settings.

Open-source controller for low-cost and high-speed atomic force microscopy imaging of skin corneocyte nanotextures.

High-speed atomic force microscopes (HS-AFMs) with high temporal resolution enable dynamic phenomena to be visualized at nanoscale resolution. However, HS-AFMs are more complex and costlier than conventional AFMs, and particulars of an open-source HS-AFM controller have not been published before. These high entry barriers hinder the popularization of HS-AFMs in both academic and industrial applications. In addition, HS-AFMs generally have a small imaging area that limits the fields of implementation. This study presents an open-source controller that enables a low-cost simplified AFM to achieve a maximum tip-sample velocity of 5,093 µm/s (9.3 s/frame, 512 × 512 pixels), which is nearly 100 times higher than that of the original controller. Moreover, the proposed controller doubles the imaging area to 46.3 × 46.3 µm2 compared to that of the original system. The low-cost HS-AFM can successfully assess the severity of atopic dermatitis (AD) by measuring the nanotexture of human skin corneocytes in constant height DC mode. The open-source controller-based HS-AFM system costs less than $4,000, which provides resource-limited research institutes with affordable access to high-throughput nanoscale imaging to further expand the HS-AFM research community.

Methotrexate Detection in Serum at Clinically Relevant Levels with Electrochemically Assisted SERS on a Benchtop, Custom Built Raman Spectrometer.

Therapeutic drug monitoring (TDM) is an essential clinical practice for optimizing drug dosing, thereby preventing adverse effects of drugs with a narrow therapeutic window, slow clearance, or high interperson pharmacokinetic variability. Monitoring methotrexate (MTX) during high-dose MTX (HD-MTX) therapy is necessary to avoid potentially fatal side effects caused by delayed elimination. Despite the efficacy of HD-MTX treatment, its clinical application in resource-limited settings is constrained due to the relatively high cost and time of analysis with conventional analysis methods. In this work, we developed (i) an electrochemically assisted surface-enhanced Raman spectroscopy (SERS) method for detecting MTX in human serum at a clinically relevant concentration range and (ii) a benchtop, Raman detection system with an integrated potentiostat, software, and data analysis unit that enables mapping of small areas of SERS substrates and quantitative SERS-based analysis. In the assay, by promoting electrostatic attraction between gold-coated nanopillar SERS substrates and MTX molecules in aqueous samples, a detection limit of 0.13 µM with a linear range of 0.43-2 µM was achieved in PBS. The implemented sample cleanup through gel filtration proved to be highly effective, resulting in a similar detection limit (0.55 µM) and linear range (1.81-5 µM) for both PBS and serum. The developed and optimized assay could also be used on the in-house built, Raman device. We showed that MTX detection can be carried out in less than 30 min with the Raman device, paving the way toward the TDM of MTX at the point-of-need and in resource-limited environments.

Low-cost, open-source XYZ nanopositioner for high-precision analytical applications.

Nanoscale positioning has numerous applications in both academia and industry. A growing number of applications require devices with long working distances and nanoscale resolutions. Friction-inertia piezoelectric positioners, which are based on the stick-slip mechanism, achieve both nanometer resolution and centimeter-scale travel. However, the requirements of complex preload mechanism, precision machining, and precise assembly increase the cost of conventional friction-inertia nanopositioners. Herein we present the design of an open-source XYZ-axis nanopositioning system. Utilizing a magnet-based stick-slip driving mechanism, the proposed XYZ nanopositioner provides several advantages, including sub-nanometer resolution, a payload capacity of up to 12 kg (horizontal), compact size, low cost, and easy assembly; furthermore, the system is adjustment-free. The performance tests validate the precision of the system in both scanning and stepping operation modes. Moreover, the resonant spectra affirm the rigidity and dynamic response of the mechanism. In addition, we demonstrate the practical applications of this nanopositioner in various measurement techniques, including scanning electron microscopy, vibrometry, and atomic force microscopy. Furthermore, we present 11 variations of the nanopositioner designs that are either compatible with ultra-high-vacuum systems and other existing systems, 3D printable, or hacking commercial linear slides.

Fast and quantitative 2D and 3D orientation mapping using Raman microscopy.

Non-destructive orientation mapping is an important characterization tool in materials science and geoscience for understanding and/or improving material properties based on their grain structure. Confocal Raman microscopy is a powerful non-destructive technique for chemical mapping of organic and inorganic materials. Here we demonstrate orientation mapping by means of Polarized Raman Microscopy (PRM). While the concept that PRM is sensitive to orientation changes is known, to our knowledge, an actual quantitative orientation mapping has never been presented before. Using a concept of ambiguity-free orientation determination analysis, we present fast and quantitative single-acquisition Raman-based orientation mapping by simultaneous registration of multiple Raman scattering spectra obtained at different polarizations. We demonstrate applications of this approach for two- and three-dimensional orientation mapping of a multigrain semiconductor, a pharmaceutical tablet formulation and a polycrystalline sapphire sample. This technique can potentially move traditional X-ray and electron diffraction type experiments into conventional optical laboratories.

Imaging of dehydration in particulate matter using Raman line-focus microscopy.

Crystalline solids can incorporate water molecules into their crystal lattice causing a dramatic impact on their properties. This explains the increasing interest in understanding the dehydration pathways of these solids. However, the classical thermal analytical techniques cannot spatially resolve the dehydration pathway of organic hydrates at the single particle level. We have developed a new method for imaging the dehydration of organic hydrates using Raman line-focus microscopy during heating of a particle. Based on this approach, we propose a new metastable intermediate of theophylline monohydrate during the three-step dehydration process of this system and further, we visualize the complex nature of the three-step dehydration pathway of nitrofurantoin monohydrate to its stable anhydrous form. A Raman line-focus mapping option was applied for fast simultaneous mapping of differently sized and shaped particles of nitrofurantoin monohydrate, revealing the appearance of multiple solid-state forms and the non-uniformity of this particle system during the complex dehydration process. This method provides an in-depth understanding of phase transformations and can be used to explain practical industrial challenges related to variations in the quality of particulate materials.

Where Is the Drug? Quantitative 3D Distribution Analyses of Confined Drug-Loaded Polymer Matrices.

To enhance oral bioavailability of poorly soluble drugs, microfabricated devices can be utilized. One example of such devices is microcontainers. These are cylindrical in shape with an inner cavity for drug loading and with only the top side open for release. Supercritical CO2 (scCO2) impregnation is an interesting technique for loading drugs into polymeric matrices in, for example, microcontainers since it avoids the use of organic solvents and is cheap. One of the main drawbacks of this technique is the unknown three-dimensional drug distribution in the polymer matrix. The aim of this study was to investigate the loading of two poorly soluble drugs, naproxen and ketoprofen, by scCO2 impregnation into confined polymer matrices of different sizes. Three different sizes of microcontainers (small, medium, and large) and, thereby, different surface areas accessible for impregnation were compared. From in vitro studies, the amount of naproxen and ketoprofen loaded into the different microcontainers and their corresponding release profiles were seen to be similar. A custom-made Raman microscope facilitated volumetric Raman maps of an entire microcontainer filled with polyvinylpyrrolidone (PVP) and scCO2 impregnated with either naproxen or ketoprofen. In all microcontainer sizes, the drugs were only detected in the top layer of the polymer matrix, explaining the observed similar release profiles. Using X-ray powder diffraction and Raman spectroscopy, the solid state form of the drugs was evaluated, showing that ketoprofen was amorphous in all microcontainer sizes. Naproxen was found not to be crystalline nor amorphous but in a less ordered configuration than the crystalline state. In conclusion, volumetric Raman mapping is a powerful technology for imaging drug distribution and drug crystallinity in polymers and allowed us to conclude that (i) scCO2 impregnation depth does not depend on surface area and (ii) impregnated drugs are noncrystalline.