Predictive value of PD-L1 and TMB for short-term efficacy prognosis in non-small cell lung cancer and construction of prediction models.

Objective: To investigate the correlation between programmed death ligand 1(PD-L1), tumor mutation burden (TMB) and the short-term efficacy and clinical characteristics of anti-PD-1 immune checkpoint inhibitor combination chemotherapy in NSCLC patients. The efficacy of the prediction model was evaluated. Methods: A total of 220 NSCLC patients receiving first-line treatment with anti-PD-1 immune checkpoint inhibitor combined with chemotherapy were retrospectively collected. The primary endpoint was short-term efficacy ORR. The correlation between short-term efficacy, PD-L1, TMB, and clinical characteristics using χ2 test or t-test was evaluated. Screen the independent prognostic factors using univariate and multivariate logistic regression analyses, and construct a nomogram prediction model using the "rms" package in R software. Using receiver operating characteristic (ROC) curve analysis to evaluate the independent Prognostic factors and the prediction model. Using decision curve analysis (DCA) to verify the superiority of the prediction model. Results: The mean values of PD-L1, TMB, neutrophils, lymphocytes, neutrophil-to-lymphocyte ratio, and albumin were the highest in the ORR group, PD-L1 expression and TMB correlated with epidermal growth factor receptor expression. Multivariate analyses showed that PD-L1, TMB, and neutrophil were independent prognostic factors for ORR. The area under the ROC curve (AUC) values of the ROC constructed based on these three indicators were 0.7104, 0.7139, and 0.7131, respectively. The AUC value under the ROC of the nomogram model was 0.813. The DCA of the model showed that all three indicators used together to build the prediction model of the net return were higher than those of the single indicator prediction model. Conclusion: PD-L1, TMB, and neutrophils are independent prognostic factors for short-term efficacy. The nomogram prediction model constructed using these three indicators can further improve predictive efficacy of ICIs in patients with NSCLC.

nELISA: A high-throughput, high-plex platform enables quantitative profiling of the secretome.

We present the nELISA, a high-throughput, high-fidelity, and high-plex protein profiling platform. DNA oligonucleotides are used to pre-assemble antibody pairs on spectrally encoded microparticles and perform displacement-mediated detection. Spatial separation between non-cognate antibodies prevents the rise of reagent-driven cross-reactivity, while read-out is performed cost-efficiently and at high-throughput using flow cytometry. We assembled an inflammatory panel of 191 targets that were multiplexed without cross-reactivity or impact on performance vs 1-plex signals, with sensitivities as low as 0.1pg/mL and measurements spanning 7 orders of magnitude. We then performed a large-scale secretome perturbation screen of peripheral blood mononuclear cells (PBMCs), with cytokines as both perturbagens and read-outs, measuring 7,392 samples and generating ~1.5M protein datapoints in under a week, a significant advance in throughput compared to other highly multiplexed immunoassays. We uncovered 447 significant cytokine responses, including multiple putatively novel ones, that were conserved across donors and stimulation conditions. We also validated the nELISA's use in phenotypic screening, and propose its application to drug discovery.

Facile Synthesis of ZnO/WO3 Nanocomposite Porous Films for High-Performance Gas Sensing of Multiple VOCs.

Volatile organic compounds (VOCs) in indoor environments have typical features of multiple components, high concentration, and long duration. The development of gas sensors with high sensitivity to multiple VOCs is of great significance to protect human health. Herein, we proposed a sensitive ZnO/WO3 composite chemi-resistive sensor facilely fabricated via a sacrificial template approach. Based on the transferable properties of self-assembled monolayer colloidal crystal (MCC) templates, two-dimensional honeycomb-like ordered porous ZnO/WO3 sensing matrixes were constructed in situ on commercial ceramic tube substrates with curved and rough surfaces. The nanocomposite thin films are about 250 nm in thickness with large-scale structural consistency and integrity, which facilitates characteristic responses with highly sensitivity and reliability. Furthermore, the nanocomposite sensor shows simultaneous responses to multiple VOCs that commonly exist in daily life with an obvious suppression sensing for traditional flammable gases. Particularly, a detection limit of 0.1 ppm with a second-level response/recovery time can be achieved, which is beneficial for real-time air quality assessments. We proposed a heterojunction-induced sensing enhancement mechanism for the ZnO/WO3 nanocomposite film in which the formation of abundant heterojunctions between ZnO and WO3 NPs significantly increases the thickness of the electron depletion layer in the bulk film and improves the formation of active oxygen species on the surface, which is conducive to enhanced responses for reducing VOC gases. This work not only provides a simple approach for the fabrication of high-performance gas sensors but also opens an achievable avenue for air quality assessment based on VOC concentration detection.

Parallel DNA circuits by autocatalytic strand displacement and nanopore readout.

DNA nanotechnology provides a unique opportunity for molecular computation, with strand displacement reactions enabling controllable reorganization of nanostructures. Additional DNA strand exchange strategies with high selectivity for input will enable novel complex systems including biosensing applications. Herein, we propose an autocatalytic strand displacement (ACSD) circuit: initiated by DNA breathing and accelerated by a seesaw catalytic reaction, ACSD ensures that only the correct base sequence starts the catalytic cycle. Analogous to an electronic circuit with a variable resistor, two ACSD reactions with different rates are connected in parallel to mimic a parallel circuit containing branches with different resistances. Finally, we introduce a multiplexed nanopore sensing platform to report the output results of a parallel path selection system at the single-molecule level. By combining the ACSD strategy with fast and sensitive single-molecule nanopore readout, a new generation of DNA-based computing tools is established.

Fast and Selective Detection of Trace Chemical Warfare Agents Enabled by an ESIPT-Based Fluorescent Film Sensor.

Reliable detection of airborne chemical warfare agents (CWAs) at the site and in real-time remains a challenge due to the rarity of miniaturized analytical tools. Herein, an o-carborane-functionalized benzothiazole derivative (PCBO) with excited-state intramolecular proton transfer (ESIPT) and AIE characteristics was synthesized. The PCBO-based film sensor showed a highly sensitive response to representative simulants of CWAs, and detection limits were found to be 1.0 mg·m-3 for triphosgene, 6.0 mg·m-3 for chloroethyl ethyl sulfide, and 0.2 mg·m-3 for diethyl chlorophosphite. Moreover, the sensor showed great reusability (>100 cycles) and unprecedented response speed (<0.5 s). The excellent sensing performance was ascribed to the microenvironmental sensitivity of the sensing fluorophore, the porous adlayer structure of the film, and the specific binding of the fluorophore to the analytes. Furthermore, discrimination and identification of the examined CWA simulants were realized via the introduction of another fluorophore (HCBO)-based film. Importantly, a portable fluorescent CWA detector was built with the sensor as the key component, and its applicability was demonstrated by the successful detection of a typical CWA sample (Sarin). The present study indicates that fluorescent film sensors could satisfy reliable onsite and real-time detection of harmful chemicals.

High-Density-Nanotips-Composed 3D Hierarchical Au/CuS Hybrids for Sensitive, Signal-Reproducible, and Substrate-Recyclable SERS Detection.

Surface-enhanced Raman scattering (SERS) provides an unprecedented opportunity for fingerprinting identification and trace-level detection in chemistry, biomedicine, materials, and so on. Although great efforts have been devoted to fabricating sensitive plasmonic nanomaterials, it is still challenging to batch-produce a SERS substrate with high sensitivity, good reproducibility, and perfect recyclability. Here, we describe a facile fabrication of three-dimensional (3D) hierarchical Au/CuS nanocomposites, in which high-density Au nanotips enable highly SERS-active sensing, and the well-defined microflower (MF) geometry produces perfect signal reproducibility (RSD < 5%) for large laser spot excitations (>50 µm2), which is particularly suitable for practical on-site detection with a handheld Raman spectrometer. In addition, a self-cleaning ability of this Au/CuS Schottky junction photocatalyst under sunlight irradiation allows complete removal of the adsorbed analytes, realizing perfect regeneration of the SERS substrates over many cycles. The mass-production, ultra-sensitive, high-reproducibility, and fast-recyclability features of hierarchical Au/CuS MFs greatly facilitate cost-effective and field SERS detection of trace analytes in practice.

Flexible and Transparent Oligothiophene-o-Carborane-Containing Hybrid Films for Nonlinear Optical Limiting Based on Efficient Two-Photon Absorption.

Structure-property relationship for fluorophores with favorable nonlinear optical (NLO) properties are promising topics in organic chemistry and material science. Herein, a series of terthiophene-o-carborane dyads and triads covalently linked with different end-capping styles were readily synthesized and comprehensively investigated. Quantitative values of the crystal and packing structures, photophysical parameters including aggregation-induced emission (AIE) and two-photon absorption (2PA) were provided. Significant impact of carborane unit for introducing the AIE characteristic has been investigated in contrast to the parent oligothiophene. All the obtained fluorophores exhibit maximum absorption around 370 nm in THF and emit bright reddish photoluminscence with absolute fluorescence quantum yields above 16% in solid states. Intramolecular charge communication between oligothiophene and carborane plays important roles in the related NLO properties. These results are supported well by the time-dependent DFT theoretical calculations. Effective 2PA cross sections (δ2PA = 95-355 GM@650 nm) and transition dipole moments of the derivatives are variable for different end-capping styles. Their potential applications as optical limiting materials based on the 2PA mechanism in solutions and doped PDMS films were further evaluated. Taken together, this work provides an understanding of their structure-property relationship, and flexible PDMS films as outstanding candidates for practical applications in optical limiting.

Rapid detection of sulfur mustard hydrolysis products based on microextraction by packed sorbent combined with nano-electrospray ionization mass spectrometry.

RATIONALE: Sulfur mustard is a blister agent prohibited by the Chemical Weapons Convention, and the detection of its hydrolysis product, thiodiglycol (TDG), is an important indicator of blister agent contamination. Due to the poor volatility and low extraction efficiency of TDG, derivatization gas chromatography or liquid chromatography is required for conventional methods, and the detection process is cumbersome and time-consuming. METHODS: A microextraction by packed sorbent (MEPS) device and a nano-electrospray ionization (nano-ESI) device were used. The central composite design (CCD) model of Response Surface Methodology was used to optimize the elution procedure; the variance analysis under equal repeated trials with multiple factors was used to quantitatively analyze the significance of the impact of related factors on the nano-ESI efficiency. The MEPS-nano-ESI-MS experimental conditions were optimized. RESULTS: A new detection method of sulfur mustard hydrolysis products in water based on MEPS-nano-ESI-MS was established; the detection limit was 1 ng/mL and was linear between 5 ng/mL and 100 ng/mL (R2 = 0.9911) with a precision of ≤7.2%, and the recovery rate was 107.89% when the sample concentration was 40 ng/mL. CONCLUSIONS: The experimental results showed that the proposed method could quickly detect the contaminated water samples without chromatographic separation and derivatization, thereby verifying the contamination of sulfur mustard on site.

High-Performance Trichloroacetic Acid Sensor Based on the Intramolecular Hydrogen Bond Formation and Disruption of a Specially Designed Fluorescent o-Carborane Derivative in the Film State.

Discriminative and sensitive detection of environmentally important and health-related trichloroacetic acid (TCA) suffers from various problems such as bulky instruments and time-consuming operation as well as complex sample processing. Herein, we present a rapid, sensitive, and specific method for the detection of gaseous TCA using a fluorescent single-molecule array. An o-carborane-based benzothiazole derivative (CB-BT-OCH3) with specific fluorescence properties was specifically designed and utilized to fabricate a film-based single-molecule array. It was revealed that the fluorescent film is photochemically stable and extremely sensitive to TCA vapor, depicting an observable fluorescence color change from green to blue. The experimental detection limit is 0.2 ppm, which is lower than the safety limit (1 ppm) required by the threshold limit values and biological exposure indices. In addition, the film could show detectable intensity change within 0.2 s. On the basis of multiple signal responses, a conceptual two-channel-based fluorescent TCA sensor was developed. Importantly, the proposed conceptual sensor paves a new route to the development of specific fluorescent film-based sensor arrays with a single fluorophore as sensing units.

The ionization process of chemical warfare agent simulants in low temperature plasma ionization.

The application of low-temperature plasma ionization technology in the chemical warfare agent detection was mostly focused on the research of rapid detection methods. Limited studies are available on the ionization process of chemical warfare agents in low temperature plasma. Through the intensity of protonated molecules of dimethyl methylphosphonate (DMMP) in different solvents including methanol, deuterated methanol (methanol-D4), pure water, and deuterium oxide (water-D2), it was concluded that the water molecule in the air provides the hydrogen ion (H+) needed for ionization. The product ion spectra and the collision-induced dissociation processes of protonated molecules of nerve agent simulants, including DMMP, diethyl methanephosphonate (DEMP), trimethyl phosphate (TMP), triethyl phosphate (TEP), tripropyl phosphate (TPP), and tributyl phosphate (TBP) were analyzed. Results revealed that H+ mostly combined with phosphorus oxygen double bond (P = O) in the low-temperature plasma ionization. By analyzing the peak intensity distribution of product ions of protonated molecules, the presence of proton and charge migration in the low temperature plasma ionization and collision-induced dissociation were researched. This study could provide technical guidance for the rapid and accurate detection of chemical warfare agents through low temperature plasma ionization-mass spectrometry.

Digital Data Storage Using DNA Nanostructures and Solid-State Nanopores.

Solid-state nanopores are powerful tools for reading the three-dimensional shape of molecules, allowing for the translation of molecular structure information into electric signals. Here, we show a high-resolution integrated nanopore system for identifying DNA nanostructures that has the capability of distinguishing attached short DNA hairpins with only a stem length difference of 8 bp along a DNA double strand named the DNA carrier. Using our platform, we can read up to 112 DNA hairpins with a separating distance of 114 bp attached on a DNA carrier that carries digital information. Our encoding strategy allows for the creation of a library of molecules with a size of up to 5 × 1033 (2112) that is only built from a few hundred types of base molecules for data storage and has the potential to be extended by linking multiple DNA carriers. Our platform provides a nanopore- and DNA nanostructure-based data storage method with convenient access and the potential for miniature-scale integration.

Importance evaluation of spectral lines in Laser-induced breakdown spectroscopy for classification of pathogenic bacteria.

The correct classification of pathogenic bacteria is significant for clinical diagnosis and treatment. Compared with the use of whole spectral data, using feature lines as the inputs of the classification model can improve the correct classification rate (CCR) and reduce the analyzing time. In order to select feature lines, we need to investigate the contribution to the CCR of each spectral line. In this paper, two algorithms, important weights based on principal component analysis (IW-PCA) and random forests (RF), were proposed to evaluate the importance of spectra lines. The laser-induced plasma spectra (LIBS) of six common clinical pathogenic bacteria species were measured and a support vector machine (SVM) classifier was used to classify the LIBS of bacteria species. In the proposed IW-PCA algorithm, the product of the loading of each line and the variance of the corresponding principal component were calculated. The maximum product of each line calculated from the first three PCs was used to represent the line's importance weight. In the RF algorithm, the Gini index reduction value of each line was considered as the line's importance weight. The experimental results demonstrated that the lines with high importance were more suitable for classification and can be chosen as feature lines. The optimal number of feature lines used in the SVM classifier can be determined by comparing the CCRs with a different number of feature lines. Importance weights evaluated by RF are more suitable for extracting feature lines using LIBS combined with an SVM classification mechanism than those evaluated by IW-PCA. Furthermore, the two methods mutually verified the importance of selected lines and the lines evaluated important by both IW-PCA and RF contributed more to the CCR.

Gold nanoparticles-enhanced ion-transmission mass spectrometry for highly sensitive detection of chemical warfare agent simulants.

Gold nanoparticles (AuNPs)-embedded paper was coupled with ion-transmission mass spectrometry (MS) to enable the highly sensitive detection of chemical warfare agent (CWA) simulants in solutions. With the assistance of a low-temperature plasma (LTP) probe, we found that AuNPs were capable to enhance the ionization efficiencies of target analytes, with MS signal intensities surprisingly undergone an 800-fold increase under optimized conditions. The interaction between AuNPs and the radiofrequency electromagnetic field was believed to promote the desorption/ionization process, resulting in the unusual signal enhancement phenomenon. Based on this finding, we established a method for the rapid analysis of two simulants of nerve agents, dimethyl methylphosphonate (DMMP) and diisopropyl methylphosphonate (DIMP), with a dynamic range from 0.5 ng/mL to 100 ng/mL and detection limits of 0.1 ng/mL and 0.3 ng/mL, respectively. As sample pretreatments have been eliminated, the developed strategy is particularly promising for the on-site detection of CWAs considering its simple and rapid analytical workflow.

Combining Affinity Selection and Specific Ion Mobility for Microchip Protein Sensing.

The sensitive detection of proteins is a key objective in many areas of biomolecular science, ranging from biophysics to diagnostics. However, sensing in complex biological fluids is hindered by nonspecific interactions with off-target species. Here, we describe and demonstrate an assay that utilizes both the chemical and physical properties of the target species to achieve high selectivity in a manner not possible by chemical complementarity alone, in complex media. We achieve this objective through a combinatorial strategy, by simultaneously exploiting free-flow electrophoresis for target selection, on the basis of electrophoretic mobility, and conventional affinity-based selection. In addition, we demonstrate amplification of the resultant signal by a catalytic DNA nanocircuit. This approach brings together the inherent solution-phase advantages of microfluidic sample handling with isothermal, enzyme-free signal amplification. With this method, no surface immobilization or washing steps are required, and our assay is well suited to monoepitopic targets, presenting advantages over conventional ELISA techniques.

Direction- and Salt-Dependent Ionic Current Signatures for DNA Sensing with Asymmetric Nanopores.

Solid-state nanopores are promising tools for single-molecule detection of both DNA and proteins. In this study, we investigated the patterns of ionic current blockades as DNA translocates into or out of the geometric confinement of conically shaped pores across a wide range of salt conditions. We studied how the geometry of a nanopore affects the detected ionic current signal of a translocating DNA molecule over a wide range of salt concentration. The blockade level in the ionic current depends on the translocation direction at a high salt concentration, and at lower salt concentrations we find a nonintuitive ionic current decrease and increase within each single event for the DNA translocations exiting from confinement. We use a recently developed method for synthesizing DNA molecules with multiple position markers, which provides further experimental characterization by matching the position of the DNA in the pore with the observed ionic current signal. Finally, we employ finite element calculations to explain the shapes of the signals observed at all salt concentrations and show that the unexpected current decrease and increase are due to the competing effects of ion concentration polarization and geometric exclusion of ions. Our analysis shows that over a wide range of geometries, voltages, and salt concentrations, we are able to understand the ionic current signals of DNA in asymmetric nanopores, enabling signal optimization in molecular sensing applications.

Single molecule based SNP detection using designed DNA carriers and solid-state nanopores.

Single nucleotide polymorphisms (SNPs) play a crucial role as molecular markers in medical and diagnostic application. We demonstrate a nanopore based method for SNP detection at the single molecule level. Designed DNA carriers are used to distinguish DNA strands containing only one single base difference and follow strand displacement kinetics.

Quantifying Nanomolar Protein Concentrations Using Designed DNA Carriers and Solid-State Nanopores.

Designed "DNA carriers" have been proposed as a new method for nanopore based specific protein detection. In this system, target protein molecules bind to a long DNA strand at a defined position creating a second level transient current drop against the background DNA translocation. Here, we demonstrate the ability of this system to quantify protein concentrations in the nanomolar range. After incubation with target protein at different concentrations, the fraction of DNA translocations showing a secondary current spike allows for the quantification of the corresponding protein concentration. For our proof-of-principle experiments we use two standard binding systems, biotin-streptavidin and digoxigenin-antidigoxigenin, that allow for measurements of the concentration down to the low nanomolar range. The results demonstrate the potential for a novel quantitative and specific protein detection scheme using the DNA carrier method.

Ionic conductivity, structural deformation, and programmable anisotropy of DNA origami in electric field.

The DNA origami technique can enable functionalization of inorganic structures for single-molecule electric current recordings. Experiments have shown that several layers of DNA molecules, a DNA origami plate, placed on top of a solid-state nanopore is permeable to ions. Here, we report a comprehensive characterization of the ionic conductivity of DNA origami plates by means of all-atom molecular dynamics (MD) simulations and nanocapillary electric current recordings. Using the MD method, we characterize the ionic conductivity of several origami constructs, revealing the local distribution of ions, the distribution of the electrostatic potential and contribution of different molecular species to the current. The simulations determine the dependence of the ionic conductivity on the applied voltage, the number of DNA layers, the nucleotide content and the lattice type of the plates. We demonstrate that increasing the concentration of Mg(2+) ions makes the origami plates more compact, reducing their conductivity. The conductance of a DNA origami plate on top of a solid-state nanopore is determined by the two competing effects: bending of the DNA origami plate that reduces the current and separation of the DNA origami layers that increases the current. The latter is produced by the electro-osmotic flow and is reversible at the time scale of a hundred nanoseconds. The conductance of a DNA origami object is found to depend on its orientation, reaching maximum when the electric field aligns with the direction of the DNA helices. Our work demonstrates feasibility of programming the electrical properties of a self-assembled nanoscale object using DNA.

Silicon nitride nanopores for nanoparticle sensing.

In recent years, nanopore has attracted broad attention as an essential technique for DNA sequencing and nanoparticle sensing. This work presented a fundamental study of nanoparticle translocation through silicon nitride nanopores. Since particles with rather high charge to mass ratio, such as DNA and proteins, have been widely studied, there was still little information about the translocation behavior of nanoparticles with low charge/mass ratio. However, these nanoparticles include viruses and colloids are important potential sample for nanopore sensing technique. In order to screen nanoparticles' size, concentration, surface property by apply silicon nitride nanopores, and fulfill specific recognition which could be applied in laboratory medicine and environmental monitoring, optimal experimental conditions needed to be confirmed. In this paper, polystyrene (PS) bead was used as an example to provide reference towards this condition. The effects of nanopore/nanoparticle diameter ratio and bias voltage on sensing results were analyzed in this work. We found that lower precision accuracy might be demonstrated by the larger pore (120 nm) while the translocation velocity tended to be slowed down. By increasing the bias voltage could favor the detection productiveness. In spite of the intense interest, the baseline fluctuations and vague event bounds aggravated are far from understood.

Voltage-driven translocation of DNA through a high throughput conical solid-state nanopore.

Nanopores have become an important tool for molecule detection at single molecular level. With the development of fabrication technology, synthesized solid-state membranes are promising candidate substrates in respect of their exceptional robustness and controllable size and shape. Here, a 30-60 (tip-base) nm conical nanopore fabricated in 100 nm thick silicon nitride (Si(3)N(4)) membrane by focused ion beam (FIB) has been employed for the analysis of λ-DNA translocations at different voltage biases from 200 to 450 mV. The distributions of translocation time and current blockage, as well as the events frequencies as a function of voltage are investigated. Similar to previously published work, the presence and configurations of λ-DNA molecules are characterized, also, we find that greater applied voltages markedly increase the events rate, and stretch the coiled λ-DNA molecules into linear form. However, compared to 6-30 nm ultrathin solid-state nanopores, a threshold voltage of 181 mV is found to be necessary to drive DNA molecules through the nanopore due to conical shape and length of the pore. The speed is slowed down ∼5 times, while the capture radius is ∼2 fold larger. The results show that the large nanopore in thick membrane with an improved stability and throughput also has the ability to detect the molecules at a single molecular level, as well as slows down the velocity of molecules passing through the pore. This work will provide more motivations for the development of nanopores as a Multi-functional sensor for a wide range of biopolymers and nano materials.