Probe Platform Research Articles

Templated Pluripotent Stem Cell Differentiation via Substratum-Guided Artificial Signaling.

The emerging field of synthetic morphogenesis implements synthetic biology tools to investigate the minimal cellular processes sufficient for orchestrating key developmental events. As the field continues to grow, there is a need for new tools that enable scientists to uncover nuances in the molecular mechanisms driving cell fate patterning that emerge during morphogenesis. Here, we present a platform that combines cell engineering with biomaterial design to potentiate artificial signaling in pluripotent stem cells (PSCs). This platform, referred to as PSC-MATRIX, extends the use of programmable biomaterials to PSCs competent to activate morphogen production through orthogonal signaling, giving rise to the opportunity to probe developmental events by initiating morphogenetic programs in a spatially constrained manner through non-native signaling channels. We show that the PSC-MATRIX platform enables temporal and spatial control of transgene expression in response to bulk, soluble inputs in synthetic Notch (synNotch)-engineered human PSCs for an extended culture of up to 11 days. Furthermore, we used PSC-MATRIX to regulate multiple differentiation events via material-mediated artificial signaling in engineered PSCs using the orthogonal ligand green fluorescent protein, highlighting the potential of this platform for probing and guiding fate acquisition. Overall, this platform offers a synthetic approach to interrogate the molecular mechanisms driving PSC differentiation that could be applied to a variety of differentiation protocols.

Realizing Orthogonal Modes in Compact Cavity‐Backed Dual‐Polarized Antenna Through Simple Feeding Structures for Millimeter‐Wave AiP Applications

AbstractThis paper presents a dual‐polarized cavity‐backed antenna designed for mm‐wave applications, featuring simple feeding structures with a high‐isolation for 60 GHz compact AiP applications. The dual‐polarization design relies on two separate feed ports that excite two orthogonal modes within the same resonant cavity, achieving very high port isolation of up to 40 dB over the entire band. We conducted a detail analysis of the antenna, including its working principles and parametric studies. For verification, we fabricated an antenna test kit using standard printed process on substrates and measured the kit from the back‐side of a GSG probing platform. The proposed antenna demonstrates a wide impedance bandwidth, stable radiation patterns, very low cross‐polarization levels, and a high radiation efficiency. The co‐located cavity‐backed design ensures the compactness and facilitates easy integration with ICs in a very small AiP module. These features make the proposed antenna highly suitable for 60 GHz AiP applications, such as high‐data‐rate wireless communication and mmW polarimetric radar systems.

A smartphone-integrated ratiometric fluorescent sensor for ascorbic acid determination using microplasma-enabled carbon dots and rhodamine B

A switchable ratiometric fluorescent smartphone-assisted sensing platform based on nitrogen-doped carbon dots (N-CDs) and Rhodamine B was fabricated for the determination of the ascorbic acid (AA) content in fruits by quenching the fluorescence of N-CDs with Hg2+ (turn-off) and recovering with AA (turn-on). The blue-emission N-CDs was synthesized by liquid dielectric barrier discharge microplasma with an average size of 3.65 nm and an absolute quantum yield of 18 % (excited at 345 nm). In addition, the fluorescence color was converted to RGB values, enabling visual and quantitative determination of AA. Under optimal parameters, the linear ranges for detecting AA were found to be 3–170 μM and 5–170 μM for fluorescence spectrometer and smartphone sensing platform. The detection limits were 0.98 μM and 2.90 μM, respectively. Furthermore, the satisfactory recoveries in fruits were obtained by RF probe and smartphone platform. This smartphone-assisted platform will facilitate sensitive and visual determination for AA.

Optical fiber SPR probe platform combined with oriented antibody optimized modification for ultrasensitive and portable detection of human thyroglobulin

Thyroglobulin (Tg) is an essential indicator for monitoring differentiated thyroid cancer. However, there are still significant challenges in achieving fast, accurate, and efficient point-of-care testing (POCT). This paper presented an optical fiber bioprobe platform technology based on the ultrasensitive sensing principle of surface plasmon resonance (SPR), aiming to achieve an efficient and precise immunoassay of Tg. The fiber probes were modified with Tg antibodies to form fiber Tg probes. After optimization of the oriented antibody modification, the detection sensitivity of fiber Tg probes increased by 3 times, facilitating one-step direct detection of μg-level sample concentrations such as thyroid tissue fluid samples. For ng-level sample concentrations such as blood samples, a two-step measurement method was used to amplify the surface load signal. Through Tg detection studies in different concentration ranges and background environments, our fiber Tg probes exhibited a sensitivity of 81.99 pm / (ng / mL) and a limit of detection of 1.4 ng / mL, and were able to complete ng-level detection in serum backgrounds. The fiber Tg probe holds immense potential in the diagnosis and postoperative monitoring of thyroid cancer. Additionally, being a miniature, disposable, reliable, and portable quantitative optical immunoassay tool, it can serve as a rapid detection instrument for various target proteins and pathogens by simply altering the modified antibody.

Dual signal amplification sensor based on imprint-modified DNA for high-sensitivity detection of copper ions

In order to improve the sensitivity of ion imprinted electrochemical (IIE) sensor to detect copper ions, a novel dual signal amplification (DSA) sensor was proposed using deoxyribonucleic acid (DNA) modified IIE sensor. Chitosan (CS) was used as a functional monomer for IIE sensor. The IIE sensor was prepared by mixing copper ion templates with CS and removing the templates. Gold nanoparticles (AuNPs) were electrodeposited onto the surface of IIE sensor to improve their conductivity and provide a connecting platform for DNA probes. And then the DNA probe was fixed on the surface of AuNPs to prepare a DSA sensor. The signal of DSA sensor was significantly amplified by the synergistic effect of IIE sensor and DNA probe. The feasibility, parameter optimization and electrochemical performance were analyzed by experimental comparison, surface micro scanning and kinetic study. The results show that compared with IIE sensor, the sensitivity of DSA sensor is significantly improved, with a value of 1.6712 × 10-6 A·(µM)-1. The exchange current density is 5.9765 × 10-8 A·cm−2. The detection range is 10 nM to 100 nM (R2 = 0.99651) with the detection limit of 4.17 nM (3σ/slope). The recommended method has high sensitivity and selectivity, as well as very high recovery rates in tap water and milk.

Superluminality in parity-time symmetric Bragg gratings

Parity-time ( PT ) symmetric Bragg gratings (PTBGs) possess unique features compared to traditional ones. For example, the photonic bandgap of a PTBG can be modified and even closed when the PT symmetry evolves from an exact phase to a broken one, and the complex reflection coefficient of a PTBG is sensitive to the direction of incidence. In this article, we reveal how the superluminal effects of transmission behave following the modified band structure of PTBGs. The superluminality of the directionally sensitive reflection is also discussed. We then investigate the Hartman effect and argue that, to account for the superluminal effects in PTBGs, a directionally sensitive dwell time should be applied. This study offers unique insights into the mechanisms of superluminality and group delay of light in non-Hermitian open systems and contributes to the advancement of PTBGs, which can eventually become an indispensable platform for probing some of the exotic properties of optical wave phenomena and light–matter interactions.

A firm-push-to-open and light-push-to-lock strategy for a general chemical platform to develop activatable dual-modality NIR-II probes.

Activatable near-infrared (NIR) imaging in the NIR-II range is crucial for deep tissue bioanalyte tracking. However, designing such probes remains challenging due to the limited availability of general chemical strategies. Here, we introduced a foundational platform for activatable probes, using analyte-triggered smart modulation of the π-conjugation system of a NIR-II-emitting rhodamine hybrid. By tuning the nucleophilicity of the ortho-carboxy moiety, we achieved an electronic effect termed "firm-push-to-open and light-push-to-lock," which enables complete spirocyclization of the probe before sensing and allows for efficient zwitterion formation when the light-pushing aniline carbamate trigger is transformed into a firm-pushing aniline. This platform produces dual-modality NIR-II imaging probes with ~50-fold fluorogenic and activatable photoacoustic signals in live mice, surpassing reported probes with generally below 10-fold activatable signals. Demonstrating generality, we successfully designed probes for hydrogen peroxide (H2O2) and hydrogen sulfide (H2S). We envision a widespread adoption of the chemical platform for designing activatable NIR-II probes across diverse applications.

A Self-Accelerating Naphthalimide-Based Probe Coupled with Upconversion Nanoparticles for Ultra-Accurate Tri-Mode Visualization of Hydrogen Peroxide.

The design and development of ultra-accurate probe is of great significance to chemical sensing in complex practical scenarios. Here, a self-accelerating naphthalimide-based probe with fast response and high sensitivity toward hydrogen peroxide (H2O2) is designed. By coupling with the specially selected upconversion nanoparticles (UCNPs), an ultra-accurate colorimetric-fluorescent-upconversion luminescence (UCL) tri-mode platform is constructed. Owing to the promoted reaction process, this platform demonstrates rapid response (< 1s), an ultra-low detection limit (4.34nM), and superb anti-interferent ability even in presence of > 21 types of oxidants, explosives, metallic salts, daily compounds, colorful or fluorescent substances. In addition, the effectiveness of this design is further verified by a sponge-based sensing chip loaded with the UCNPs/probe in recognizing trace H2O2 vapor from interferents with the three characteristic colors existing simultaneously. The proposed design of probe and tri-mode visualization detection platform is expected to open up a brand-new methodology for ultra-accurate sensing.

Identification of activity-based biomarkers for early-stage pancreatic tumors in blood using single-molecule enzyme activity screening

Single-molecule enzyme activity-based enzyme profiling (SEAP) is a methodology to globally analyze protein functions in living samples at the single-molecule level. It has been previously applied to detect functional alterations in phosphatases and glycosidases. Here, we expand the potential for activity-based biomarker discovery by developing a semi-automated synthesis platform for fluorogenic probes that can detect various peptidases and protease activities at the single-molecule level. The peptidase/protease probes were prepared on the basis of a 7-amino-4-methylcoumarin fluorophore. The introduction of a phosphonic acid to the core scaffold made the probe suitable for use in a microdevice-based assay, while phosphonic acid served as the handle for the affinity separation of the probe using Phos-tag. Using this semi-automated scheme, 48 fluorogenic probes for the single-molecule peptidase/protease activity analysis were prepared. Activity-based screening using blood samples revealed altered single-molecule activity profiles of CD13 and DPP4 in blood samples of patients with early-stage pancreatic tumors. The study shows the power of single-molecule enzyme activity screening to discover biomarkers on the basis of the functional alterations of proteins.

Modular Design Platform for Activatable Fluorescence Probes Targeting Carboxypeptidases Based on ProTide Chemistry.

Carboxypeptidases (CPs) are a family of hydrolases that cleave one or more amino acids from the C-terminal of peptides or proteins and play indispensable roles in various physiological and pathological processes. However, only a few highly activatable fluorescence probes for CPs have been reported, and there is a need for a flexibly tunable molecular design platform to afford a range of fluorescence probes for CPs for biological and medical research. Here, we focused on the unique activation mechanism of ProTide-based prodrugs and established a modular design platform for CP-targeting florescence probes based on ProTide chemistry. In this design, probe properties such as fluorescence emission wavelength, reactivity/stability, and target CP can be readily tuned and optimized by changing the four probe modules: the fluorophore, the substituent on the phosphorus atom, the linker amino acid at the P1 position, and the substrate amino acid at the P1' position. In particular, switching the linker amino acid at position P1 enabled us to precisely optimize the reactivity for target CPs. As a proof-of-concept, we constructed probes for carboxypeptidase M (CPM) and prostate-specific membrane antigen (also known as glutamate carboxypeptidase II). The developed probes were applicable for the imaging of CP activities in live cells and in clinical specimens from patients. This design strategy should be useful in studying CP-related biological and pathological phenomena.

Ultrasensitive levofloxacin electrochemical biosensor based on semiconducting covalent organic framework/poly-L-cysteine/triangular Ag nanoplates modified glassy carbon electrode.

Anovel electrochemical biosensor with excellent performance was fabricated for levofloxacin (LEV) detection, which adopted triangular Ag nanoplates (Tri-AgNP) confined in a poly-L-cysteine (poly-L-Cys) film and a semiconducting covalent organic framework (COF) as the electrochemical sensing material. The developed electrochemical sensor revealed excellent analytical properties because of its good electrical conductivity, fast electron transfer, and abundant bioactive site. Based on this, a linear relationship between the LEV concentration and the peak current response at 0.92 V was obtained under the optimal experimental conditions by differential pulse voltammetry (DPV), with a wide linear range of 0.05 to 600 μM and a low limit of detection (LOD) of 0.0061 μM. The prepared sensor also realized sensitive and accurate determination of LEV in human serum and urine samples by standard addition method, with satisfactory recoveries (97.1 to 104%) and a low relative standard deviation of less than 4.6%. These results indicated that the novel ternary system has a promising application in the development of electrochemical signal probe and electrochemical biosensing platform.

Fluorescent paper sensor based on green emissive carbon nanodots for visual detection of acid orange II in food samples

This work for the first time presents a visual fluorescence paper sensor for accurate and on-site detection of acid orange II (AO2) in food matrix, where green-emissive carbon nanodots (G-CDs) were exploited. The as-synthesized G-CDs had an average particle size of 3.9 ± 0.7 nm and displayed green emission at 511 nm upon excitation at 450 nm. When there existed AO2, the G-CDs fluorescence was effectively suppressed via inner filter effect (IFE) and dynamic interaction. The as-prepared G-CDs probe achieved a 0.01–25.0 μM linear scope and a 36.5 nM limit of detection. Particularly, owing to a distinguishable reduction in G-CDs emission intensity by a smartphone application, a visual probing platform along with a nanopaper-based device were be developed for recognizing AO2. The corresponding detection limits were severally 0.61 μM and 1.39 μM. Success was achieved in applying our fluorescence approaches for detecting AO2 in Yuba, chili powder, sunflower seeds, tomato paste and orange juice samples, attaining desirable results. The present work underlines the potential of smartphone-integrated nanopaper based on CDs as an encouraging candidate for on-site AO2 recognition in food matrix.

Prenatal delta-9-tetrahydrocannabinol exposure is associated with changes in rhesus macaque DNA methylation enriched for autism genes

BackgroundWith the growing availability of cannabis and the popularization of additional routes of cannabis use beyond smoking, including edibles, the prevalence of cannabis use in pregnancy is rapidly increasing. However, the potential effects of prenatal cannabis use on fetal developmental programming remain unknown.ResultsWe designed this study to determine whether the use of edible cannabis during pregnancy is deleterious to the fetal and placental epigenome. Pregnant rhesus macaques consumed a daily edible containing either delta-9-tetrahydrocannabinol (THC) (2.5 mg/7 kg/day) or placebo. DNA methylation was measured in 5 tissues collected at cesarean delivery (placenta, lung, cerebellum, prefrontal cortex, and right ventricle of the heart) using the Illumina MethylationEPIC platform and filtering for probes previously validated in rhesus macaque. In utero exposure to THC was associated with differential methylation at 581 CpGs, with 573 (98%) identified in placenta. Loci differentially methylated with THC were enriched for candidate autism spectrum disorder (ASD) genes from the Simons Foundation Autism Research Initiative (SFARI) database in all tissues. The placenta demonstrated greatest SFARI gene enrichment, including genes differentially methylated in placentas from a prospective ASD study.ConclusionsOverall, our findings reveal that prenatal THC exposure alters placental and fetal DNA methylation at genes involved in neurobehavioral development that may influence longer-term offspring outcomes. The data from this study add to the limited existing literature to help guide patient counseling and public health polices focused on prenatal cannabis use in the future.

An optical and visual multi-mode sensing platform base on nitrogen, sulfur, boron co-doped carbon dots for rapid and simple determination of ferric ions in water

Abnormal iron ions levels may lead to some diseases and serious environmental pollution. Herein, optical and visual detection strategies of Fe3+ in water based on co-doped carbon dots (CDs) were established in the present study. Firstly, a one-pot synthetic strategy for the preparation of the N, S, B co-doped CDs with a home microwave oven was developed. Secondly, the optical properties, chemical structures, and morphology of CDs were further characterized by fluorescence spectroscopy, Uv–vis absorption spectroscopy, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and transmission electron microscope. Finally, the results indicated that the fluorescence of the co-doped CDs was quenched by ferric ions via the static mechanism and the aggregation of CDs, accompanied by the increased red color. The multi-mode sensing strategies of Fe3+ with fluorescence photometer, UV–visible spectrophotometer, portable colorimeter and smartphone had the advantages of good selectivity, excellent stability and high sensitivity. Fluorophotometry based on co-doped CDs was a powerful probe platform for measuring lower concentrations of Fe3+ due to its higher sensitivity, better linear relationship, lower limit of detection (0.27 μM) and limit of quantitation (0.91 μM). In addition, the visual detection methods with a portable colorimeter and smartphone had been proven to be very suitable for rapid and simple sensing of higher concentrations of Fe3+. Moreover, the co-doped CDs utilized for Fe3+ probes in tap water and boiler water obtained satisfactory results. Consequently, the efficient, versatile optical and visual multi-mode sensing platform could be extended to apply such a visual analysis of ferric ions in the biological, chemical and other fields.

Peptide Oxidation Induced by Liquid-Solid Contact Electrification as Revealed in Liquid Microjunction-Surface Sampling Probe Mass Spectrometry.

Reproducible peptide oxidation was observed using a homebuilt liquid microjunction-surface sampling probe (LMJ-SSP) platform for analyzing peptide standards. Although electrochemical oxidation and corona discharges have previously been associated with analyte oxidation in electrospray ionization (ESI) and ESI-related ambient ionization mass spectrometry (MS) methods, they were unlikely the causes for the peptide oxidation observed in the LMJ-SSP studies. A systematic investigation demonstrated that analyte oxidation was induced during the droplet drying on a solid surface through liquid-solid electrification processes. To minimize unwanted analyte oxidation, the water content in the sample solution should be decreased and the use of hydroxyl-functionalized substrates, such as glass slides, should be avoided. In addition, if water is an essential solvent component, adding an antioxidant, such as ascorbic acid, to the sample solution before droplet evaporation on the solid surface could lower the percentage of analyte oxidation. The present findings apply to all the MS methods that involve drying microliters of sample solution onto a suitable substrate in their sample preparation protocols.

Target-triggered enzyme-free amplification for highly efficient AND-gated bioimaging in living cells.

Rapid, simultaneous, and sensitive detection of biomolecules has important application prospects in disease diagnosis and biomedical research. However, because the content of intracellular endogenous target biomolecules is usually very low, traditional detection methods can't be used for effective detection and imaging, and to enhance the detection sensitivity, signal amplification strategies are frequently required. The hybridization chain reaction (HCR) has been used to detect many disease biomarkers because of its simple operation, good reproducibility, and no enzyme involvement. Although HCR signal amplification methods have been employed to detect and image intracellular biomolecules, there are still false positive signals. Therefore, a target-triggered enzyme-free amplification system (GHCR system) was developed, as a fluorescent AND-gated sensing platform for intracellular target probing. The false positive signals can be well avoided and the accuracy of detection and imaging can be improved by using the design of the AND gate. Two cancer markers, GSH and miR-1246, were used as two orthogonal inputs for the AND gated probe. The AND-gated probe only works when GSH and miR-1246 are the inputs at the same time, and FRET signals can be the output. In addition to the use of AND-gated imaging, FRET-based high-precision ratiometric fluorescence imaging was employed. FRET-based ratiometric fluorescent probes have a higher ability to resist interference from the intracellular environment, they can avoid false positive signals well, and they are expected to have good specificity. Due to the advantages of HCR, AND-gated, and FRET fluorescent probes, the GHCR system exhibited highly efficient AND-gated FRET bioimaging for intracellular endogenous miRNAs with a lower detection limit of 18 pM, which benefits the applications of ratiometric intracellular biosensing and bioimaging and offers a novel concept for advancing the diagnosis and therapeutic strategies in the field of cancer.

Low LOD and Non-Adsorption Probing of Functionalized Magnetic Beads Using Quartz Crystal Microbalance Under Bimodal Magnetic Field

Low limit of detection (LOD) and highly sensitive probing of functional magnetic nanobead labels is the cornerstone of magnetic label molecular detection technology, which is of great significance in analytical chemistry and biomolecular analysis. Here we present a probing technology based on quartz crystal microbalance (QCM) combines bimodal magnetic fields to accurate and rapid probing of magnetic beads. The proposed technique uses QCM to detect the change of viscosity and density of tested solution under a bimodal magnetic field, thereby enabling the quantification of magnetic bead concentration. The experimental findings demonstrate that the technique can detect magnetic beads accurately and rapidly within 0 ng/mL to 100 μg/mL without anchoring magnetic beads on the QCM. Furthermore, when the particle size of magnetic beads is 1 μm and 0.28 μm, the LOD is reached at 4.62 ng/mL and 2.31 ng/mL, respectively. The versatility of the no-modification probing platform yet the wide detection range and low LOD to various magnetic particles, showcase the potential of the proposed technology to enable swift, repeatable, and dependable assays across a broad range of applications, including but not limited to biomolecule analysis, Molecular genetics, pre-symptomatic diagnosis, and biosafety.

A novel oil-water microemulsion strategy for controllable synthesis of large mesoporous carbon nanoparticles

Recently, the carbon nanoparticles with defined architecture, small particle diameter and large mesopore size, have aroused people's enormous interests on account of their laudable properties. Here, it is novel mesoporous carbon nanoparticles (MCNs) that have been successfully fabricated in a neoteric oil-water microemulsion system, which is developed to synchronously adjust the surface roughness, particle diameter, and large mesopore size through accommodating the emulsion polarity. Besides, pentanol as a cosurfactant not only significantly affects the evolution of mesopore size but also acts an vital role in the interfacial interaction between carbon precursors and soft templates, leading to the fluffy growth of particles. Interestingly, an extended swelling-penetration mechanism is subtly put forward to explain the formation and transformation of MCNs, which would provide a simple and unique ingenuity for this material species. Remarkably, to fulfill the multifunctional properties of such MCNs, MCNs-2 is further utilized as a fluorescent probe platform to detect mutated nucleic acids with high selectivity and low detection limit, as well as a medical carrier to work well in drug delivery system with high adsorption/release properties and extremely low cytotoxicity, which would empower it with luciferous prospects for the applications in biosensor and biomedical fields.

Polymethine Molecular Platform for Ratiometric Fluorescent Probes in the Second near-Infrared Window.

Visualizing biomolecules such as enzymes in the deep tissue of living organisms via molecular ratiometric fluorescent probes in the second near-infrared window (NIR-II) with a built-in self-calibration function can provide reliable information about relevant pathophysiological processes directly but so far is not feasible due to the lack of a fluorescence modulation strategy in the NIR-II window. Here we present a molecular platform Py-2 by integrating the rhodamine 6G scaffold and polymethine. The maximal emission wavelength of Py-2 was 1010 nm and blue-shifted to 945 nm when its secondary amine was acylated. Based on Py-2, two molecular ratiometric NIR-II fluorescent probes, nitroreductase-responsive Rap-N and ROS-responsive Rap-R, were constructed and successfully demonstrated in vitro and in vivo. Overall, this report presents a unique approach to developing ratiometric NIR-II molecular probes for in vivo biosensing.

A Portable Smartphone-Based System for the Detection of Blood Calcium Using Ratiometric Fluorescent Probes.

Hypocalcemia is a disease that adversely affects the production and reproduction of dairy cows. A portable device for rapid bovine blood calcium sensing has been growing in demand. Herein, we report a smartphone-based ratiometric fluorescence probe (SRFP) platform as a new way to detect and quantify calcium ions (Ca2+) in blood serum. Specifically, we employed a cost-effective and portable smartphone-based platform coupled with customized software that evaluates the response of Ca2+ ions to ratiometric fluorescence probe in bovine serum. The platform consists of a three-dimensional (3D) printed housing and low-cost optical components that excite fluorescent probe and selectively transmit fluorescence emissions to smartphones. The customized software is equipped with a calibration model to quantify the acquired fluorescence images and quantify the concentration of Ca2+ ions. The ratio of the green channel to the red channel bears a highly reproducible relationship with Ca2+ ions concentration from 10 μM to 40 μM in bovine serum. Our detection system has a limit of detection (LOD) of 1.8 μM in bovine serum samples and the recoveries of real samples ranged from 92.8% to 110.1%, with relative standard deviation (RSD) ranging from 1.72% to 4.89%. The low-cost SRFP platform has the potential to enable campesino to rapidly detect Ca2+ ions content in bovine serum on-demand in any environmental setting.