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Metals play an essential role in biological systems and are required as structural or catalytic co-factors in many proteins. Disruption of the homeostatic control and/or spatial distributions of metals can lead to disease. Imaging technologies have been developed to visualize elemental distributions across a biological sample. Measurement of elemental distributions by imaging mass spectrometry and imaging X-ray fluorescence are increasingly employed with technologies that can assess histological features and molecular compositions. Data from several modalities can be interrogated as multimodal images to correlate morphological, elemental, and molecular properties. Elemental and molecular distributions have also been axially resolved to achieve three-dimensional volumes, dramatically increasing the biological information. In this review, we provide an overview of recent developments in the field of metal imaging with an emphasis on multimodal studies in two and three dimensions. We specifically highlight studies that present technological advancements and biological applications of how metal homeostasis affects human health.
Copyright © 2020 Elsevier Ltd. All rights reserved.
A label-free optical biosensor based on a one-dimensional photonic crystal microring resonator with enhanced light-matter interaction is demonstrated. More than a 2-fold improvement in volumetric and surface sensing sensitivity is achieved compared to conventional microring sensors. The experimental bulk detection sensitivity is ~248nm/RIU and label-free detection of DNA and proteins is reported at the nanomolar scale. With a minimum feature size greater than 100nm, the photonic crystal microring resonator biosensor can be fabricated with the same standard lithographic techniques used to mass fabricate conventional microring resonators.
A flow-through sensing platform based on open-ended porous silicon (PSi) microcavity membranes that are compatible with integration in on-chip sensor arrays is demonstrated. Because of the high aspect ratio of PSi nanopores, the performance of closed-ended PSi sensors is limited by infiltration challenges and slow sensor responses when detecting large molecules such as proteins and nucleic acids. In order to improve molecule transport efficiency and reduce sensor response time, open-ended PSi nanopore membranes were used in a flow-through sensing scheme, allowing analyte solutions to pass through the nanopores. The molecular binding kinetics in these PSi membranes were compared through experiments and simulation with those from closed-ended PSi films of comparable thickness in a conventional flow-over sensing scheme. The flow-through PSi membrane resulted in a 6-fold improvement in sensor response time when detecting a high molecular weight analyte (streptavidin) versus in the flow-over PSi approach. This work demonstrates the possibility of integrating multiple flow-through PSi sensor membranes within parallel microarrays for rapid and multiplexed label-free biosensing.
The ability of cells to adhere and sense differences in tissue stiffness is crucial for organ development and function. The central mechanisms by which adherent cells detect extracellular matrix compliance, however, are still unknown. Using two single-molecule-calibrated biosensors that allow the analysis of a previously inaccessible but physiologically highly relevant force regime in cells, we demonstrate that the integrin activator talin establishes mechanical linkages following cell adhesion, which are indispensable for cells to probe tissue stiffness. Talin linkages are exposed to a range of piconewton forces and bear, on average, 7-10 pN during cell adhesion depending on their association with F-actin and vinculin. Disruption of talin's mechanical engagement does not impair integrin activation and initial cell adhesion but prevents focal adhesion reinforcement and thus extracellular rigidity sensing. Intriguingly, talin mechanics are isoform specific so that expression of either talin-1 or talin-2 modulates extracellular rigidity sensing.
This work examines the influence of charge density and surface passivation on the DNA-induced corrosion of porous silicon (PSi) waveguides in order to improve PSi biosensor sensitivity, reliability, and reproducibility when exposed to negatively charged DNA molecules. Increasing the concentration of either DNA probes or targets enhances the corrosion process and masks binding events. While passivation of the PSi surface by oxidation and silanization is shown to diminish the corrosion rate and lead to a saturation in the changes by corrosion after about 2 h, complete mitigation can be achieved by replacing the DNA probe molecules with charge-neutral PNA probe molecules. A model to explain the DNA-induced corrosion behavior, consistent with experimental characterization of the PSi through Fourier transform infrared spectroscopy and prism coupling optical measurements, is also introduced.
Peptide nucleic acids (PNA) are a unique class of synthetic molecules that have a peptide backbone and can hybridize with nucleic acids. Here, a versatile method has been developed for the automated, in situ synthesis of PNA from a porous silicon (PSi) substrate for applications in gene therapy and biosensing. Nondestructive optical measurements were performed to monitor single base additions of PNA initiated from (3-aminopropyl)triethoxysilane attached to the surface of PSi films, and mass spectrometry was conducted to verify synthesis of the desired sequence. Comparison of in situ synthesis to postsynthesis surface conjugation of the full PNA molecules showed that surface mediated, in situ PNA synthesis increased loading 8-fold. For therapeutic proof-of-concept, controlled PNA release from PSi films was characterized in phosphate buffered saline, and PSi nanoparticles fabricated from PSi films containing in situ grown PNA complementary to micro-RNA (miR) 122 generated significant anti-miR activity in a Huh7 psiCHECK-miR122 cell line. The applicability of this platform for biosensing was also demonstrated using optical measurements that indicated selective hybridization of complementary DNA target molecules to PNA synthesized in situ on PSi films. These collective data confirm that we have established a novel PNA-PSi platform with broad utility in drug delivery and biosensing.
Simple and rapid methods for detecting mRNA biomarkers from patient samples are valuable in settings with limited access to laboratory resources. In this report, we describe the development and evaluation of a self-contained assay to extract and quantify mRNA biomarkers from complex samples using a novel nucleic acid-based molecular sensor called quadruplex priming amplification (QPA). QPA is a simple and robust isothermal nucleic acid amplification method that exploits the stability of the G-quadruplex nucleotide structure to drive spontaneous strand melting from a specific DNA template sequence. Quantification of mRNA was enabled by integrating QPA with a magnetic bead-based extraction method using an mRNA-QPA interface reagent. The assay was found to maintain >90% of the maximum signal over a 4 °C range of operational temperatures (64-68 °C). QPA had a dynamic range spanning four orders of magnitude, with a limit of detection of ~20 pM template molecules using a highly controlled heating and optical system and a limit of detection of ~250 pM using a less optimal water bath and plate reader. These results demonstrate that this integrated approach has potential as a simple and effective mRNA biomarker extraction and detection assay for use in limited resource settings.
Effective point-of-care diagnostics require a biomarker detection strategy that is low-cost and simple-to-use while achieving a clinically relevant limit of detection. Here we report a biosensor that uses secondary flows arising from surface Marangoni stresses in an evaporating drop to concentrate target-mediated particle aggregates in a visually detectable spot. The spot size increases with increasing target concentration within the dynamic range of the assay. The particle deposition patterns are visually detectable and easily measured with simple optical techniques. We use optical coherence tomography to characterize the effect of cross-sectional flow fields on the motion of particles in the presence and absence of target (aggregated and non-aggregated particles, respectively). We show that choice of substrate material and the presence of salts and glycerol in solution promote the Marangoni-induced flows that are necessary to produce signal in the proposed design. These evaporation-driven flows generate signal in the assay on a PDMS substrate but not substrates with greater thermal conductivity like indium tin oxide-coated glass. In this proof-of-concept design we use the M13K07 bacteriophage as a model target and 1 μm-diameter particles surface functionalized with anti-M13 monoclonal antibodies. Using standard microscopy-based techniques to measure the final spot size, the assay has a calculated limit-of-detection of approximately 100 fM. Approximately 80% of the maximum signal is generated within 10 minutes of depositing a 1 μL drop of reacted sample on PDMS enabling a relatively quick time-to-result.
A porous silicon (PSi) grating-coupled Bloch surface and sub-surface wave (BSW/BSSW) biosensor is demonstrated to size selectively detect the presence of both large and small molecules. The BSW is used to sense large immobilized analytes at the surface of the structure while the BSSW that is confined inside but near the top of the structure is used to sensitively detect small molecules. Functionality of the BSW and BSSW modes is theoretically described by dispersion relations, field confinements, and simulated refractive index shifts within the structure. The theoretical results are experimentally verified by detecting two different small chemical molecules and one large 40 base DNA oligonucleotide. The PSi-BSW/BSSW structure is benchmarked against current porous silicon technology and is shown to have a 6-fold higher sensitivity in detecting large molecules and a 33% improvement in detecting small molecules. This is the first report of a grating-coupled BSW biosensor and the first report of a BSSW propagating mode.
© 2013 Published by Elsevier B.V.