A tempo-spatial controllable microfluidic shear-stress generator for in-vitro mimicking of the thrombus.

Pathological conditions linked to shear stress have been identified in hematological diseases, cardiovascular diseases, and cancer. These conditions often exhibit significantly elevated shear stress levels, surpassing 1000 dyn/cm2 in severely stenotic arteries. Heightened shear stress can induce mechanical harm to endothelial cells, potentially leading to bleeding and fatal consequences. However, current technology still grapples with limitations, including inadequate flexibility in simulating bodily shear stress environments, limited range of shear stress generation, and spatial and temporal adaptability. Consequently, a comprehensive understanding of the mechanisms underlying the impact of shear stress on physiological and pathological conditions, like thrombosis, remains inadequate. To address these limitations, this study presents a microfluidic-based shear stress generation chip as a proposed solution. The chip achieves a substantial 929-fold variation in shear stress solely by adjusting the degree of constriction in branch channels after PDMS fabrication. Experiments demonstrated that a rapid increase in shear stress up to 1000 dyn/cm2 significantly detached 88.2% cells from the substrate. Long-term exposure (24 h) to shear stress levels below 8.3 dyn/cm2 did not significantly impact cell growth. Furthermore, cells exposed to shear stress levels equal to or greater than 8.3 dyn/cm2 exhibited significant alterations in aspect ratio and orientation, following a normal distribution. This microfluidic chip provides a reliable tool for investigating cellular responses to the wide-ranging shear stress existing in both physiological and pathological flow conditions.

Continuous production of bimetallic nanoparticles on carbon nanotubes based on 3D-printed microfluidics.

Nanoparticle-functionalized carbon nanotubes are promising in many research fields, especially in sensing, due to their intriguing performance in catalysis. However, these nanomaterials are mainly produced through batch processes under harsh conditions, thus encountering inherent limitations of low throughput and uncontrollable morphology of functional nanoparticles (NPs). In this work, we propose a method for high-yield and continuous production of bimetallic (Pt-Pd) NPs on multi-walled carbon nanotubes (MWCNTs) at room temperature through a custom 3D-printed microfluidic platform. A homogenous particle nucleation and growth environment could be created on the microfluidic platform that was equipped with two 3D-printed micromixers. Pt-Pd NPs loaded on MWCNTs were prepared in the microfluidic platform with high throughput and controlled size, dispersity and composition. The synthetic parameters for these nanocomposites were investigated to optimize their electrocatalytic performance. The optimized nanocomposites exhibited excellent electrocatalytic activity with exceptional sensitivity and wide detection range, superior to their counterparts prepared via conventional approaches. This method proposed here could be further adapted for manufacturing other catalyst support materials, opening more avenues for future large-scale production and catalytic investigation of functional nanomaterials.

High sensitivity and automatic chemiluminescence detection of glucose and lactate using a spin-disc paper-based device.

This paper reports a spin-disc paper-based device with 10 individual detection units containing electromagnetic modules controlling the sample incubation time before chemiluminescence (CL) signal detection. After the sample was added to the top paper chip and incubated with the enzyme, the electromagnet was turned off to allow contact between the top and bottom paper. The H2O2 generated by the sample flowed vertically to the bottom paper and initiated the oxidase of the luminol to generate the CL signal. After one detection the disc was automatically rotated to the next position to repeat the above detection. The advantage of using the device over the lateral flow and the in situ detection was firstly proved using the detection of H2O2 and the glucose/lactate sample with 5 minute incubation. The CL intensity was increased 300 times/1000 times as the glucose/lactate was incubated for 5 minutes compared to the non-incubated samples. Afterward, the device was employed to separately detect glucose and lactate diluted in PBS, artificial sweat, artificial saliva, and fresh cell culture media. Finally, the device was employed to detect the glucose and lactate in the media collected over the 24 hour culture of PC3 cells. The uptake and production rates of glucose and lactate were correspondingly determined as 0.328 ± 0.015 pmol h-1 per cell and 1.254 ± 0.053 pmol h-1 per cell, respectively. The reported device has wide application potential due to its capabilities in automatic detection of multiple samples with very high sensitivity and small sample volume (down to 0.5 µL).

Smart Droplet Microfluidic System for Single-Cell Selective Lysis and Real-Time Sorting Based on Microinjection and Image Recognition.

Single-cell analysis has important implications for understanding the specificity of cells. To analyze the specificity of rare cells in complex blood and biopsy samples, selective lysis of target single cells is pivotal but difficult. Microfluidics, particularly droplet microfluidics, has emerged as a promising tool for single-cell analysis. In this paper, we present a smart droplet microfluidic system that allows for single-cell selective lysis and real-time sorting, aided by the techniques of microinjection and image recognition. A custom program evolved from Python is proposed for recognizing target droplets and single cells, which also coordinates the operation of various parts in a whole microfluidic system. We have systematically investigated the effects of voltage and injection pressure applied to the oil-water interface on droplet microinjection. An efficient and selective droplet injection scheme with image feedback has been demonstrated, with an efficiency increased dramatically from 2.5% to about 100%. Furthermore, we have proven that the cell lysis solution can be selectively injected into target single-cell droplets. Then these droplets are shifted into the sorting area, with an efficiency for single K562 cells reaching up to 73%. The system function is finally explored by introducing complex cell samples, namely, K562 cells and HUVECs, with a success rate of 75.2% in treating K562 cells as targets. This system enables automated single-cell selective lysis without the need for manual handling and sheds new light on the cooperation with other detection techniques for a broad range of single-cell analysis.

Pollution-Free and Highly Sensitive Lactate Detection in Cell Culture Based on a Microfluidic Chip.

Cell metabolite detection is important for cell analysis. As a cellular metabolite, lactate and its detection play an important role in disease diagnosis, drug screening and clinical therapeutics. This paper reports a microfluidic chip integrated with a backflow prevention channel for cell culture and lactate detection. It can effectively realize the upstream and downstream separation of the culture chamber and the detection zone, and prevent the pollution of cells caused by the potential backflow of reagent and buffer solutions. Due to such a separation, it is possible to analyze the lactate concentration in the flow process without contamination of cells. With the information of residence time distribution of the microchannel networks and the detected time signal in the detection chamber, it is possible to calculate the lactate concentration as a function of time using the de-convolution method. We have further demonstrated the suitability of this detection method by measuring lactate production in human umbilical vein endothelial cells (HUVEC). The microfluidic chip presented here shows good stability in metabolite quick detection and can work continuously for more than a few days. It sheds new insights into pollution-free and high-sensitivity cell metabolism detection, showing broad application prospects in cell analysis, drug screening and disease diagnosis.

Development and validation of a nomogram model for predicting unfavorable functional outcomes in ischemic stroke patients after acute phase.

Introduction: Prediction of post-stroke functional outcome is important for personalized rehabilitation treatment, we aimed to develop an effective nomogram for predicting long-term unfavorable functional outcomes in ischemic stroke patients after acute phase. Methods: We retrospectively analyzed clinical data, rehabilitation data, and longitudinal follow-up data from ischemic stroke patients who underwent early rehabilitation at multiple centers in China. An unfavorable functional outcome was defined as a modified Rankin Scale (mRS) score of 3-6 at 90 days after onset. Patients were randomly allocated to either a training or test cohort in a ratio of 4:1. Univariate and multivariate logistic regression analyses were used to identify the predictors for the development of a predictive nomogram. The area under the receiver operating characteristic curve (AUC) was used to evaluate predictive ability in both the training and test cohorts. Results: A total of 856 patients (training cohort: n = 684; test cohort: n = 172) were included in this study. Among them, 518 patients experienced unfavorable outcomes 90 days after ischemic stroke. Trial of ORG 10172 in Acute Stroke Treatment classification (p = 0.024), antihypertensive agents use [odds ratio (OR) = 1.86; p = 0.041], 15-day Barthel Index score (OR = 0.930; p < 0.001) and 15-day mRS score (OR = 13.494; p < 0.001) were selected as predictors for the unfavorable outcome nomogram. The nomogram model showed good predictive performance in both the training (AUC = 0.950) and test cohorts (AUC = 0.942). Conclusion: The constructed nomogram model could be a practical tool for predicting unfavorable functional outcomes in ischemic stroke patients underwent early rehabilitation after acute phase.

Multiplexed detection of biomarkers using a microfluidic chip integrated with mass-producible micropillar array electrodes.

Quantifying multiple biomarkers with high sensitivity in tiny biological samples is essential to meet the growing demand for point-of-care testing. This paper reports the development of a novel microfluidic device integrated with mass-producible micropillar array electrodes (µAEs) for multiple biomarker detections. The µAE are mass-fabricated by soft lithography and hot embossing technique. Pt-Pd bimetallic nanoclusters (BNC) are modified on the surface of µAEs by constant potential (CP)/multi-potential step (MPS) electrodeposition strategies to improve the electroanalytical performance. The experimental result displays that Pt-Pd BNC/µAEs have good sensitivity enhancement compared with bare planar electrodes and bare µAEs, the enhancement being 56.5 and 9.5 times respectively, from the results of the H2O2 detection. Furthermore, glucose, uric acid and sarcosine were used as model biomarkers to show the biosensing capability with high sensitivity. The linear range and LOD of the glucose, uric acid and sarcosine detection are 0.1 mM-12 mM, 10 µM-800 µM and 2.5 µM-100 µM, 58.5, 3.4 and 0.4 µM, respectively. In particular, biosensing chips show wide linear ranges covering required detection ranges of glucose, uric acid and sarcosine in human serum, indicating the developed device has great potential in self-health management and clinical requirements.

Up-to-date progress in bioprinting of bone tissue.

The major apparatuses used for three-dimensional (3D) bioprinting include extrusion-based, droplet-based, and laser-based bioprinting. Numerous studies have been proposed to fabricate bioactive 3D bone tissues using different bioprinting techniques. In addition to the development of bioinks and assessment of their printability for corresponding bioprinting processes, in vitro and in vivo success of the bioprinted constructs, such as their mechanical properties, cell viability, differentiation capability, immune responses, and osseointegration, have been explored. In this review, several major considerations, challenges, and potential strategies for bone bioprinting have been deliberated, including bioprinting apparatus, biomaterials, structure design of vascularized bone constructs, cell source, differentiation factors, mechanical properties and reinforcement, hypoxic environment, and dynamic culture. In addition, up-to-date progress in bone bioprinting is summarized in detail, which uncovers the immense potential of bioprinting in re-establishing the 3D dynamic microenvironment of the native bone. This review aims to assist the researchers to gain insights into the reconstruction of clinically relevant bone tissues with appropriate mechanical properties and precisely regulated biological behaviors.

Development of a novel microfluidic biosensing platform integrating micropillar array electrode and acoustic microstreaming techniques.

Quantifying biomarkers at the early stage of the disease is challenging due to the low abundance of biomarkers in the sample and the lack of sensitive techniques. This article reports the development of a novel microfluidic electrochemical biosensing platform to address this challenge. The electrochemical sensing is achieved by utilizing a micropillar array electrode (µAE) coated with 3D bimetallic Pt-Pd nanotrees to enhance the sensitivity. A bubble-based acoustic microstreaming technique is integrated with the device to increase the contact of analyte molecules with the surface of electrodes to further enhance the electrochemical performance. The current density of Pt-Pd NTs/µAE with acoustic microstreaming is nearly 22 times that of the bare planar electrode in potassium ferrocyanide solution. The developed biosensor has demonstrated excellent sensing performance. For hydrogen peroxide detection, both the Pt-Pd NTs/µAE and acoustic microstreaming contribute to the sensitivity enhancement. The current density of the Pt-Pd NTs/µAE is approximatively 28 times that of the bare µAE. With acoustic microstreaming, this enhancement is further increased by nearly 1.6 times. The platform has a linear detection range of 5-1000 µM with a LOD of 1.8 µM toward hydrogen peroxide detection, while for sarcosine detection, the linear range is between 5 and 100 µM and LOD is 2.2 µM, respectively. Furthermore, the sarcosine biosensing shows a high sensitivity of 667 µA mM-1∙cm-2. Such a sensing platform has the potential as a portable device for high sensitivity detection of biomarkers.

Experimental Study on Capillary Microflows in High Porosity Open-Cell Metal Foams.

Metal foams have been widely used in heat pipes as wicking materials. The main issue with metal foams is the surface property capillary limit. In this paper, a chemical blackening process for creating a superhydrophilic surface on copper foams is studied with seven different NaOH and NaClO2 solution concentrations (1.5~4.5 mol/L), in which the microscopic morphology of the treated copper foam surface is analyzed by scanning electron microscopy. The capillary experiments are carried out to quantify the wicking characteristics of the treated copper foams and the results are compared with theoretical models. A the microscope is used to detect the flow stratification characteristics of the capillary rise process. The results show that the best wicking ability is obtained for the oxidation of copper foam using 3.5 mol/L of NaOH and NaClO2 solution. Gravity plays a major role in defining the permeability and effective pore radius, while the effect of evaporation can be ignored. The formation of a fluid stratified interface between the unsaturated and saturated zone results in capillary performance degradation. The current study is important for understanding the flow transport in porous materials.

High-Throughput Microfluidic Production of Bimetallic Nanoparticles on MXene Nanosheets and Application in Hydrogen Peroxide Detection.

Nanoparticle-functionalized transition-metal carbides and nitrides (MXenes) have attracted extensive attention in electrochemical detection owing to their excellent catalytic performance. However, the mainstream synthetic routes rely on the batch method requiring strict experimental conditions, generally leading to low yield and poor size tunability of particles. Herein, we report a high-throughput and continuous microfluidic platform for preparing a functional MXene (Ti3C2Tx) with bimetallic nanoparticles (Pt-Pd NPs) at room temperature. Two 3D micromixers with helical elements were integrated into the microfluidic platform to enhance the secondary flow for promoting transport and reaction in the synthesis process. The rapid mixing and strong vortices in these 3D micromixers prevent aggregation of NPs in the synthesis process, enabling a homogeneous distribution of Pt-Pd NPs. In this study, Pt-Pd NPs loaded on the MXene nanosheets were synthesized under various hydrodynamic conditions of 1-15 mL min-1 with controlled sizes, densities, and compositions. The mean size of Pt-Pd NPs could be readily controlled within the range 2.4-9.3 nm with high production rates up to 13 mg min-1. In addition, synthetic and electrochemical parameters were separately optimized to improve the electrochemical performance of Ti3C2Tx/Pt-Pd. Finally, the optimized Ti3C2Tx/Pt-Pd was used for hydrogen peroxide (H2O2) detection and shows excellent electrocatalytic activity. The electrode modified with Ti3C2Tx/Pt-Pd here presents a wide detection range for H2O2 from 1 to 12 000 µM with a limit of detection down to 0.3 µM and a sensitivity up to 300 µA mM-1 cm-2, superior to those prepared in the traditional batch method. The proposed microfluidic approach could greatly enhance the electrochemical performance of Ti3C2Tx/Pt-Pd, and sheds new light on the large-scale production and catalytic application of the functional nanocomposites.

Nomogram Prediction Model Analysis of Risk Factors for Conversion to Thoracotomy after Thoracoscopic Resection of Lung Cancer and Prognostic Value of Lung Cancer.

This study was aimed at exploring the risk factors for thoracotomy in patients undergoing thoracoscopic resection of lung cancer and further analyzing the factors affecting the prognosis of patients. Ninety-six patients with non-small-cell lung cancer who underwent thoracoscopic pulmonary resection were recruited as the subjects, and they were enrolled into the thoracoscopic group (n = 88) and the thoracotomy group (n = 8) according to whether thoracotomy was performed. Univariate analysis and logistic multivariate regression were performed to analyze the risk factors for conversion to thoracotomy, and nomogram prediction model was employed to analyze the prognostic factors. The results revealed that the proportion of patients over 65 years old, with history of coronary heart disease, diabetes, and pulmonary tuberculosis, etc., in the thoracotomy group and the thoracoscopic group was significantly different (P < 0.05). There were statistically significant differences in the development of interlobular cleft, pleural adhesion, tumor diameter > 3.5 cm, vascular and lymph node invasion, and tumor TNM stage between the thoracotomy group and the thoracoscopic group (P < 0.05). Overall, the age of patients ≥ 65 years old, tumor diameter > 3.5 cm, hypoplasia of interlobular fissure, history of pulmonary tuberculosis, pleural adhesion, and TNM stage IIIa were all independent risk factors for thoracoscopic resection of lung cancer to thoracotomy. Cox model and nomogram prediction model analysis showed that surgery methods, tumor diameter > 3.5 cm, chemotherapy cycle < 4, chemotherapy, and TNM stage IIIa were all independent factors influencing the prognosis of patients undergoing thoracoscopic lung cancer resection. This nomogram prediction model had high application value in patient prognosis prediction.

Size-Based Sorting and In Situ Clonal Expansion of Single Cells Using Microfluidics.

Separation and clonal culture and growth kinetics analysis of target cells in a mixed population is critical for pathological research, disease diagnosis, and cell therapy. However, long-term culture with time-lapse imaging of the isolated cells for clonal analysis is still challenging. This paper reports a microfluidic device with four-level filtration channels and a pneumatic microvalve for size sorting and in situ clonal culture of single cells. The valve was on top of the filtration channels and used to direct fluid flow by membrane deformation during separation and long-term culture to avoid shear-induced cell deformation. Numerical simulations were performed to evaluate the influence of device parameters affecting the pressure drop across the filtration channels. Then, a droplet model was employed to evaluate the impact of cell viscosity, cell size, and channel width on the pressure drop inducing cell deformation. Experiments showed that filtration channels with a width of 7, 10, 13, or 17 µm successfully sorted K562 cells into four different size ranges at low driving pressure. The maximum efficiency of separating K562 cells from media and whole blood was 98.6% and 89.7%, respectively. Finally, the trapped single cells were cultured in situ for 4-7 days with time-lapse imaging to obtain the lineage trees and growth curves. Then, the time to the first division, variation of cell size before and after division, and cell fusion were investigated. This proved that cells at the G1 and G2 phases were of significantly distinct sizes. The microfluidic device for size sorting and clonal expansion will be of tremendous application potential in single-cell studies.

Droplet Microfluidics Enables Tracing of Target Cells at the Single-Cell Transcriptome Resolution.

The rapid promotion of single-cell omics in various fields has begun to help solve many problems encountered in research, including precision medicine, prenatal diagnosis, and embryo development. Meanwhile, single-cell techniques are also constantly updated with increasing demand. For some specific target cells, the workflow from droplet screening to single-cell sequencing is a preferred option and should reduce the impact of operation steps, such as demulsification and cell recovery. We developed an all-in-droplet method integrating cell encapsulation, target sorting, droplet picoinjection, and single-cell transcriptome profiling on chips to achieve labor-saving monitoring of TCR-T cells. As a proof of concept, in this research, TCR-T cells were encapsulated, sorted, and performed single-cell transcriptome sequencing (scRNA-seq) by injecting reagents into droplets. It avoided the tedious operation of droplet breakage and re-encapsulation between droplet sorting and scRNA-seq. Moreover, convenient device operation will accelerate the progress of chip marketization. The strategy achieved an excellent recovery performance of single-cell transcriptome with a median gene number over 4000 and a cross-contamination rate of 8.2 ± 2%. Furthermore, this strategy allows us to develop a device with high integrability to monitor infused TCR-T cells, which will promote the development of adoptive T cell immunotherapy and their clinical application.

Numerical Study on a Bio-Inspired Micropillar Array Electrode in a Microfluidic Device.

The micropillar array electrode (µAE) has been widely applied in microchip-based electrochemical detection systems due to a large current response. However, it was found that amplifying the current through further adjusting geometrical parameters is generally hindered by the shielding effect. To solve this problem, a bio-inspired micropillar array electrode (bµAE) based on the microfluidic device has been proposed in this study. The inspiration is drawn from the structure of leatherback sea turtles' mouths. By deforming a µAE to rearrange the micropillars on bilateral sides of the microchannel, the contact area between micropillars and analytes increases, and thus the current is substantially improved. A numerical simulation was then used to characterize the electrochemical performance of bµAEs. The effects of geometrical and hydrodynamic parameters on the current of bµAEs were investigated. Moreover, a prototypical microchip integrated with bµAE was fabricated for detailed electrochemical measurement. The chronoamperometry measurements were conducted to verify the theoretical performance of bµAEs, and the results suggest that the experimental data are in good agreement with those of the simulation model. This work presents a novel bµAE with great potential for highly sensitive electrochemical detection and provides a new perspective on the efficient configuration of the µAE.

A peptide translated from circPPP1R12A promotes the malignancy of non-small cell lung cancer cells through AKT signaling pathway.

BACKGROUND: Recent literature have indicated that the malignancy of cancer cells is modulated by hsa_circ_0000423 (named circPPP1R12A) through the way of translating protein. Herein, we investigated the role and latent mechanisms of circPPP1R12A in Non-Small Cell Lung Cancer (NSCLC). METHODS: CircPPP1R12A expression was measured by qRT-PCR. The malignancy of NSCLC was determined by CCK-8, TUNEL assay, Wound healing, Transwell and Western blotting assays. The underlying mechanisms of circPPP1R12A were confirmed by Western blotting and qRT-PCR assays. RESULTS: CircPPP1R12A expression in NSCLC tissues was higher than that of neighboring normal tissues. CircPPP1R12A showed an upregulated expression in NSCLC cells. Upregulation of circPPP1R12A could promote the cell viability of NSCLC cells and reduce the apoptosis of NSCLC cells, while it could not promote cell invasion and migration. The reduction of cell viability and apoptosis was discovered in NSCLC cells with the silencing of circPPP1R12A, but circPPP1R12A knockdown does not inhibit cell invasion and migration. There was something interesting that circPPP1R12A encoding protein circPPP1R12A-73aa was found in NSCLC cells. Mutations in circPPP1R12a-73AA might disrupt the function of circPPP1ra-73AA in A549 and H1299 cells. Next, we found that circPPP1R12A caused the increased growth of NSCLC cells by activating AKT signaling pathway. CONCLUSION: In summary, our study proved that NSCLC cell proliferation was promoted by circPPP1R12A-73aa translated from circPPP1R12A through the AKT pathway, which could throw some light on the understanding of the mechanism of NSCLC.

Microflows in two-generation alveolar cells at an acinar bifurcation.

The alveolus is a basic functional unit of the human respiratory system, and the airflow in the alveoli plays an important role in determining the transport and deposition of particulate matter, which is crucial for inhaled disease diagnosis and drug delivery. In the present study, taking advantage of the precise control ability of the microfluidic technique, a rhythmically expanding alveolar chip with multiple alveoli in two generations is designed and both the geometric and kinematic similarities are matched with the real human respiration system. With the help of a micro-PIV measurement system, the microflow patterns inside each alveolus can be studied. The observed vortex and radial flow patterns and the discovery of stagnant saddle points are similar to those captured in our previous platform with only one alveolus [Lv et al., Lab Chip 20, 2394-2402 (2020)]. However, the interactions between multiple alveoli also uncover new phenomena, such as the finding of stagnant saddle points in non-vortex flow patterns and significant differences in the flow pattern around the points between the time of T/4 and 3T/4. The obtained results could enrich the understanding of microflow in a whole alveolar tree with multiple generations.

[Short term clinical observation of cervical anterior Hybrid surgery].

OBJECTIVE: To investigate the short-term clinical effect of the cervical anterior Hybrid surgery in the treatment of two-segment and three-segment cervical spondylosis. METHODS: From January 2018 to January 2019, 108 patients who were performed anterior Hybrid surgery with cervical degenerative diseases were collected. The patients were divided into a two-segment group with 52 patients and a three-segment group with 56 patients according to surgical segments. In two-segment group, there were 24 males and 28 females, aged from 35 to 67 years old with an average of(45.94±14.67) years old. In three-segment group, there were 23 males and 33 females, aged from 32 to 65 years old with an average of (47.54±15.34) years old. The outcome indicators of the two groups were compared. Clinical indicators:neck disability index(NDI) was used to evaluate daily life ability, Japanese Orthopedic Association(JOA) score was used to evaluate neurological function improvement, visual analogue scale(VAS) was used to evaluate pain intensity, and general clinical results were graded according to Odom's score. Cervical range of motion (ROM), fusion and complications were measured by X-ray, CT and MRI. RESULTS: All operations were successfully completed and all patients were followed up for more than 12 months. The operation time of two-segment group and three-segment group were 95 to 180 min with an average of(152.30±44.74) min and 110 to 210 min with an average of (165.18±45.86) mins, the blood loss were 20 to 100 ml with an average of (32.88±8.75) ml and 20 to 150 ml with an average of(34.64±10.63) ml respectively which has no statistical differences between the two groups (P>0.05). Compared with those before surgery, NDI, JOA, VAS and Odom's scores between two groups were significantly improved at 12 months after operation(P<0.05). However, there was no significant difference in the NDI, JOA and Odom's scores between two groups (P>0.05), and VAS in three-segment group was higher than that in two-segment group. There was no significant difference in C3-C7 cervical mobility between two groups. Surgical incisions healed smoothly in all patients without complication such as spinal cord injury and cerebrospinal fluid leakage. The bone fusion of the two groups were 43 cases (82.69%) and 45 cases(80.35%) respectively. In two-segment group, there were 2 cases of adjacent segmental hyperosteogeny, and there were 3 cases of adjacent segmental hyperosteogeny and 1 case of adjacent posterior longitudinal ligament ossification in the three-segment group. In addition, in three-segment group, there was 1 case of looseness of implants with no obvious clinical symptoms. CONCLUSION: The anterior Hybrid surgery in treating multi-level cervical spondylosis could not only improve clinical symptoms of patients but also preserve mobility. Meanwhile, the efficacy and safety of Hybrid surgery in different multi-level cervical disc diseases are confirmed, proving its value in clinical practice.

A Ti3 C2 TX /Pt-Pd based amperometric biosensor for sensitive cancer biomarker detection.

The detection of cancer biomarkers is of great significance for the early screening of cancer. Detecting the content of sarcosine in blood or urine has been considered to provide a basis for the diagnosis of prostate cancer. However, it still lacks simple, high-precision and wide-ranging sarcosine detection methods. In this work, a Ti3 C2 TX /Pt-Pd nanocomposite with high stability and excellent electrochemical performance has been synthesized by a facile one-step alcohol reduction and then used on a glassy carbon electrode (GCE) with sarcosine oxidase (SOx ) to form a sarcosine biosensor (GCE/Ti3 C2 TX /Pt-Pd/SOx ). The prominent electrocatalytic activity and biocompatibility of Ti3 C2 TX /Pt-Pd enable the SOx to be highly active and sensitive to sarcosine. Under the optimized conditions, the prepared biosensor has a wide linear detection range to sarcosine from 1 to 1000 µM with a low limit of detection of 0.16 µM (S/N = 3) and a sensitivity of 84.1 µA/mM cm2 . Besides, the reliable response in serum samples shows its potential in the early diagnosis of prostate cancer. More importantly, the successful construction and application of the amperometric biosensor based on Ti3 C2 TX /Pt-Pd will provide a meaningful reference for detecting other cancer biomarkers.

Generation and Screening of Antigen-Specific Nanobodies from Mammalian Cells Expressing the BCR Repertoire Library Using Droplet-Based Microfluidics.

Nanobodies, also known as VHHs, originate from the serum of Camelidae. Nanobodies have considerable advantages over conventional antibodies, including smaller size, more modifiable, and deeper tissue penetration, making them promising tools for immunotherapy and antibody-drug development. A high-throughput nanobody screening platform is critical to the rapid development of nanobodies. To date, droplet-based microfluidic systems have exhibited improved performance compared to the traditional phage display technology in terms of time and throughput. In realistic situations, however, it is difficult to directly apply the technology to the screening of nanobodies. Requirements of plasma cell enrichment and high cell viability, as well as a lack of related commercial reagents, are leading causes for impeding the development of novel methods. We overcame these obstacles by constructing a eukaryotic display system that secretes nanobodies utilizing homologous recombination and eukaryotic transformation technologies, and the significant advantages are that it is independent of primary cell viability and it does not require plasma cell enrichment in advance. Next, a signal capture system of "SA-beads + Biotin-antigen + nanobody-6 × His + fluorescence-labeled anti-6 × His (secondary antibody)" was designed for precise localization of the eukaryotic-expressed nanobodies in a droplet. Based on this innovation, we screened 293T cells expressing anti-PD-L1 nanobodies with a high positive rate of targeted cells (up to 99.8%). Then, single-cell transcriptomic profiling uncovered the intercellular heterogeneity and BCR sequence of target cells at a single-cell level. The complete complementarity determining region (CDR3) structure was obtained, which was totally consistent with the BCR reference. This study expanded the linkage between microfluidic technology and nanobody applications and also showed potential to accelerate the rapid transformation of nanobodies in the large-scale market.