High Manganese Content of Lipid NanoMn (LNM) by Microfluidic Technology for Enhancing Anti-Tumor Immunity.

Immunotherapy is a clinically effective method for treating tumors. Manganese can activate the cGAS-STING signaling pathway and induce an anti-tumor immune response. However, its efficacy is hindered by non-specific distribution and low uptake rates. In this study, we employed microfluidic technology to design and develop an innovative preparation process, resulting in the creation of a novel manganese lipid nanoparticle (LNM). The lipid manganese nanoparticle produced in this process boasts a high manganese payload, excellent stability, the capacity for large-scale production, and high batch repeatability. LNM has effectively demonstrated the ability to activate the cGAS-STING signaling pathway, induce the production of pro-inflammatory cytokines, and inhibit tumor development. Notably, LNM does not require combination chemotherapy drugs or other immune activators. Therefore, LNM presents a safe, straightforward, and efficient strategy for anti-tumor immune activation, with the potential for scalable production.

Tumor-Associated Monocytes Reprogram CD8+ T Cells into Central Memory-Like Cells with Potent Antitumor Effects.

CD8+ T cells are critical for host antitumor responses, whereas persistent antigenic stimulation and excessive inflammatory signals lead to T cell dysfunction or exhaustion. Increasing early memory T cells can improve T cell persistence and empower T cell-mediated tumor eradication, especially for adoptive cancer immunotherapy. Here, it is reported that tumor-associated monocytes (TAMos) are highly correlated with the accumulation of CD8+ memory T cells in human cancers. Further analysis identifies that TAMos selectively reprogram CD8+ T cells into T central memory-like (TCM-like) cells with enhanced recall responses. L-NMMA, a pan nitric oxide synthase inhibitor, can mitigate TAMo-mediated inhibition of T cell proliferation without affecting TCM-like cell generation. Moreover, the modified T cells by TAMo exposure and L-NMMA treatment exhibit long-term persistence and elicit superior antitumor effects in vivo. Mechanistically, the transmembrane protein CD300LG is involved in TAMo-mediated TCM-like cell polarization in a cell-cell contact-dependent manner. Thus, the terminally differentiated TAMo subset (CD300LGhighACElow) mainly contributes to TCM-like cell development. Taken together, these findings establish the significance of TAMos in boosting T-cell antitumor immunity.

DEAR model in overweight endometrial cancer patients undergoing fertility-sparing treatment: A randomized controlled trial.

OBJECTIVE: To evaluate the effects of DEAR weight management in overweight patients undergoing fertility-sparing treatment for endometrial cancer or atypical hyperplasia. METHODS: Women with endometrial cancer or atypical hyperplasia who received fertility-sparing treatment and had a body mass index of >25 kg/m2 were randomly allocated to the DEAR (DEAR weight management) and control (self weight management) groups. Body morphology and composition, glycolipid metabolism, and tumor outcomes were assessed in both groups before and at 3 and 6 months after intervention. RESULTS: Overall, 72 subjects were included (36 in each group). Following intervention, the DEAR group showed significantly lower median body weight (69.45 vs. 78.05), body mass index (26.19 vs. 29.15), lipid accumulation index (29.21 vs. 57.86), body fat mass (24.00 vs. 29.30), visceral fat area (112.5 vs. 133.3), and glycolipid metabolic indices (except high density lipoprotein) than the control group (P < 0.05) and showed a decreasing trend. The test group achieved significantly higher complete remission (88.46% vs. 57.14%; P < 0.05); the time to complete remission did not differ significantly (P > 0.05). CONCLUSIONS: DEAR weight management can improve the studied parameters and complete remission rates in this population. REGISTRATION: NCT06169449.

Insula Connectivity Abnormalities Predict Impulsivity in Chronic Heroin Use Disorder: A Cross-Sectional Resting-State fMRI Study.

Patients with heroin use disorder (HUD) often exhibit trait impulsivity, which may be an important factor in and a good predictor of addiction. However, the factor structure of HUD trait impulsivity (motor, attentional, and nonplanning) and its neural correlates are not yet known. A total of 24 male volunteers with HUD and 16 healthy control volunteers were recruited for this cross-sectional study. The Barratt Impulsiveness Scale (BIS-11) and resting-state functional magnetic resonance imaging (rs-fMRI) were employed using the insula as a seed point in an effort to understand the association between trait impulsivity and its intrinsic factors and functional connectivity (FC) between the insula and the whole brain. The HUD group in this study exhibited higher total trait impulsivity scores, motor impulsivity, and nonplanning impulsivity than the control group. Changes in FC between the right insula and the lateral occipital cortex and the right angular gyrus were significantly positively correlated with total trait impulsivity scores, motor impulsivity, and nonplanning impulsivity, whereas changes in the FC between the left insula and the left superior frontal gyrus and left frontopolar brain region were significantly negatively correlated with trait impulsivity. Thus, the insula may serve as an important biomarker for identifying trait impulsivity and its intrinsic factor structure in patients with HUDs.

Proline uptake promotes activation of lymphoid tissue inducer cells to maintain gut homeostasis.

Metabolic regulation is integral to the proper functioning of innate lymphoid cells, yet the underlying mechanisms remain elusive. Here, we show that disruption of exogenous proline uptake, either through dietary restriction or by deficiency of the proline transporter Slc6a7, in lymphoid tissue inducer (LTi) cells, impairs LTi activation and aggravates dextran sodium sulfate-induced colitis in mice. With an integrative transcriptomic and metabolomic analysis, we profile the metabolic characteristics of various innate lymphoid cell subsets and reveal a notable enrichment of proline metabolism in LTi cells. Mechanistically, defective proline uptake diminishes the generation of reactive oxygen species, previously known to facilitate LTi activation. Additionally, LTi cells deficient in Slc6a7 display downregulation of Cebpb and Kdm6b, resulting in compromised transcriptional and epigenetic regulation of interleukin-22. Furthermore, our study uncovers the therapeutic potential of proline supplementation in alleviating colitis. Therefore, these findings shed light on the role of proline in facilitating LTi activation and ultimately contributing to gut homeostasis.

Reliability and validity of the Chinese post-discharge coping difficulty scale-parent form in parents of premature infants: a multicenter cross-sectional study.

Background: The measurement of the coping difficulties of parents of premature infants after discharge provides objective data for nurses to prepare infants for discharge. However, no Chinese scale has been developed to measure parents' coping difficulties after their premature infants are discharged. Aim: To translate the parent version of the Post-Discharge Coping Difficulty Scale (Ped-PDCDS) from English to Chinese and test the reliability and validity of the Chinese version in parents of premature infants. Methods: A multicenter cross-sectional study of 356 parents of premature infants was conducted. The scale was symmetrically translated. Validity was evaluated in terms of content, construct, discriminant, and convergent validities. Reliability was assessed in terms of internal consistency, split-half reliability, and test-retest reliability. Results: The Chinese Ped-PDCDS finally contained 11 items. Exploratory and confirmatory factor analyses results showed that the Chinese Ped-PDCDS had three dimensions, and the convergent and discriminant validities of the scale was satisfactory. The overall reliability, split-half reliability, and test-retest reliability of the scale was 0.85, 0.92, and 0.84, respectively. Conclusion: The Chinese Ped-PDCDS has adequate psychometric properties, and is an easy and appropriate instrument for measuring parents' difficulty in coping with premature infants.

Regulation of antigen-specific T cell infiltration and spatial architecture in multiple myeloma and premalignancy.

Entry of antigen-specific T cells into human tumors is critical for immunotherapy, but the underlying mechanisms are poorly understood. Here, we combined high-dimensional spatial analyses with in vitro and in vivo modeling to study the mechanisms underlying immune infiltration in human multiple myeloma (MM) and its precursor monoclonal gammopathy of undetermined significance (MGUS). Clustered tumor growth was a feature of MM but not MGUS biopsies, and this growth pattern was reproduced in humanized mouse models. MM biopsies exhibited intralesional as well as spatial heterogeneity, with coexistence of T cell-rich and T cell-sparse regions and the presence of areas of T cell exclusion. In vitro studies demonstrated that T cell entry into MM clusters was regulated by agonistic signals and CD2-CD58 interactions. Upon adoptive transfer, antigen-specific T cells localized to the tumor site but required in situ DC-mediated antigen presentation for tumor entry. C-type lectin domain family 9 member A-positive (CLEC9A+) DCs appeared to mark portals of entry for gradients of T cell infiltration in MM biopsies, and their proximity to T cell factor 1-positive (TCF1+) T cells correlated with disease state and risk status. These data illustrate a role for tumor-associated DCs and in situ activation in promoting the infiltration of antigen-specific T cells in MM and provide insights into spatial alterations in tumor/immune cells with malignant evolution.

ACE2 inhibits proliferation of smooth muscle cell through AT1R and its downstream signaling pathway.

To investigate the effect of the angiotensin converting enzyme 2 (ACE2) on AT1R expression, ERK1/2 and STAT3 protein phosphorylation in rat vascular smooth muscle cells (VSMCs) was studied. VSMCs were transfected with a lentiviral vector including the ACE2 gene and with siRNA to regulate the level of ACE2 in VSMCs. The levels of mRNA and proteins of ACE2, AT1R, ERK1/2, p-ERK1/2, STAT3, and p-STAT3 in VSMCs were examined using real-time PCR and western blot. The proliferation of VSMCs was observed by CCK-8 assay and BrdU measurement. Upregulation of ACE2 inhibited the growth of cells elicited by angiotensin II (Ang II). ACE2 significantly suppressed the level of the AT1 receptor (AT1R) protein induced by Ang II and phosphorylated the ERK1/2 and STAT3 proteins in the downstream signaling pathway. The transcriptional and translational levels of ACE2 were significantly lower in the si-ACE2 group than in the control group. The level of AT1R mRNA and protein, both with the phosphorylation expression of ERK1/2 and STAT3 protein in the siACE2 group and the Ang II group, were significantly enhanced than those in the control group. ACE2 significantly inhibited the growth of VSMCs. ACE2 inhibited the proliferation of VSMCs by suppressing AT1R and the downstream ERK1/2 and STAT3 signaling axes. Also, Ang II enhanced the level of AT1R and phosphorylated ERK1/2 and STAT3 by inhibiting the level of the ACE2 mRNA and protein.

Genetic distinction between functional tissue-resident and conventional natural killer cells.

Tissue-residential natural killer (trNK) cells act as pioneering responders during infectious challenges. However, their discrimination with conventional NK (cNK) cells is still an issue. Through an integrative transcriptome comparison of the two NK subgroups from different tissues, we have defined two genesets capable of efficiently distinguishing them. Based on the two genesets, a fundamental difference between the activation of trNK and cNK is identified and further confirmed. Mechanistically, we have discovered a particular role of chromatin landscape in regulating the trNK activation. In addition, IL-21R and IL-18R are respectively highly expressed by trNK and cNK, indicating a role of cytokine milieu in determining their differential activation. Indeed, IL-21 is particularly critical in accessorily promoting trNK activation using a bunch of bifunctional transcription factors. Together, this study sheds light on the bona fide difference between trNK and cNK, which will further expand our knowledge about their distinct functionalities during immune responses.

Differentiating environmental scenarios to establish geochemical baseline values for heavy metals in soil: A case study of Hainan Island, China.

Soil heavy metal distributions exhibit regional heterogeneity due to the complex characteristics of parent materials and soil formation processes, emphasizing the need for appropriate regional standards prior to assessing soil risks. This study focuses on Hainan Island and employs the Multi-purpose Regional Geochemical Survey dataset to establish heavy metal geochemical baseline and background values for soil using an iterative method. Geographical detector analysis reveals that parent materials are the primary factor influencing heavy metal distribution, followed by soil types and land use. Heavy metal geochemical baseline values are established for the island's three environments and administrative regions. Notably, a universal geochemical baseline value cannot adequately represent regional variations in heavy metal distribution, with parent materials playing a crucial role in various scenarios. Locally applicable values based on parent material are the most representative for Hainan Island. This study provides a reference framework for developing region-specific environmental baseline values for soil heavy metal assessments.

Development of a DNA aptamer targeting IDO1 with anti-tumor effects.

Immune checkpoint blockade has become an effective approach to reverse the immune tolerance of tumor cells. Indoleamine 2,3-dioxygenase 1 (IDO1) is frequently upregulated in many types of cancers and contributes to the establishment of an immunosuppressive cancer microenvironment, which has been thought to be a potential target for cancer therapy. However, the development of IDO1 inhibitors for clinical application is still limited. Here, we isolated a DNA aptamer with a strong affinity and inhibitory activity against IDO1, designated as IDO-APT. By conjugating with nanoparticles, in situ injection of IDO-APT to CT26 tumor-bearing mice significantly suppresses the activity of regulatory T cells and promotes the function of CD8+ T cells, leading to tumor suppression and prolonged survival. Therefore, this functional IDO1-specific aptamer with potent anti-tumor effects may serve as a potential therapeutic strategy in cancer immunotherapy. Our data provide an alternative way to target IDO1 in addition to small molecule inhibitors.

Effects of monoacylglycerols with different saturation degrees on physical and whipping properties of milk fat-based whipping creams.

Milk fat-based whipping cream is primarily comprised of cream and whole milk. It has melt-in-the-mouth texture and unique milk flavor. However, milk fat-based whipping cream suffers from poor emulsion stability and foam firmness. The effects of monoacylglycerols (MAGs) with different saturation degrees (M1: 98% saturation, M2: 70% saturation and M3: 30% saturation) on emulsion properties (average particle size, viscosity, and emulsion stability) and whipping properties (overrun, firmness, shape retention ability, and foam stability) of milk fat-based whipping creams were investigated in this study. MAGs significantly decreased particle sizes (from 2.84 to 1.16 µm) and enhanced viscosity (from 350 to 490 cP) of the milk fat-based emulsions (emulsion without MAGs: M0, 5.01 µm, 298 cP) (P < 0.05). MAGs increased the stability of the milk fat-based emulsions with lesser phase separation during centrifugation tests and lower changes in particle sizes and viscosities during temperature cycling tests. Emulsion M1 with highest degree of saturation is less likely to destabilize and phase inverse. The decrease sharply in conductivity can be attributed to the entrapment of large amounts of air. Following that, the conductivity of M1 with low variation indicating high whipping resistance and less likely to coalescence and phase separation. Adding MAGs can significantly enhance overrun (M1: 205.3%, M2: 198.5%, M3: 141.4%) as compared to the control sample (M0: 97.9%) (P < 0.05). In emulsions containing MAGs with high degree of saturation (M1 and M2), firmness (M1: 95 g, M2: 109 g) and shape retention ability of the whipped creams were reduced as compared to control emulsion without MAG (M0: 173 g), but the foam stability (M1: 89%, M2: 91%) was enhanced (M0: 81%); M3 (firmness: 507 g; foam stability: 66%) has the contrasted effects. Whipping cream M2 demonstrated the best whipping properties with high overrun (198.46%), good firmness (109 g), shape retention ability and foam stability (91%). Good quality whipping creams can be obtained by selecting suitable MAGs.

Effects of recorded maternal voice on anthropometric parameters and heart rate in premature infants: A pilot randomised controlled trial.

PURPOSE: To explore the benefits of a recorded maternal voice intervention on weight, recumbent length, head circumference, and heart rate of preterm infants in the neonatal intensive care unit. METHODS: A pilot randomised controlled trial was conducted in this study. Preterm infants in the neonatal intensive care unit (N = 109) were recruited and randomly assigned to an intervention or control group. Both groups received routine nursing care, while preterm infants in the intervention group received a recorded maternal voice program of 20 min, twice daily for 21 days. Preterm infants' daily weight, recumbent length, head circumference, and heart rate were collected during the 21-day intervention. Participants' heart rate in the intervention group was also recorded once a day pre-during-after the recorded maternal voice program. RESULTS: Preterm infants in the intervention group showed a significant increase in weight (-75.94, 95% CI -108.04, -43.85, P < 0.001), recumbent length (-0.54, 95% CI -0.76, -0.32, P < 0.001), and head circumference (-0.37, 95%CI -0.56, -0.18, P < 0.001) compared with the control group. Preterm infants in the intervention group also showed significant changes in heart rate pre-during-after the recorded maternal voice program. However, no significant differences were found in the heart rate scores between the two groups. DISCUSSION: The changes in heart rate pre-during-after the intervention may help explain participants' more significant increase in weight, recumbent length, and head circumference. PRACTICE IMPLICATIONS: The recorded maternal voice intervention could be incorporated into clinical practice to promote growth and development in preterm infants in the neonatal intensive care unit. STUDY REGISTRATION: Australian New Zealand Clinical Trials Register, https://www.anzctr.org.au/; (registration number: ACTRN12622000019707).

Structural and biochemical insights into the molecular mechanism of TRPT1 for nucleic acid ADP-ribosylation.

Nucleic acid ADP-ribosylation has been established as a novel modification found in a wide diversity of prokaryotic and eukaryotic organisms. tRNA 2'-phosphotransferase 1 (TRPT1/TPT1/KptA) possesses ADP-ribosyltransferase (ART) activity and is able to ADP-ribosylate nucleic acids. However, the underlying molecular mechanism remains elusive. Here, we determined crystal structures of TRPT1s in complex with NAD+ from Homo sapiens, Mus musculus and Saccharomyces cerevisiae. Our results revealed that the eukaryotic TRPT1s adopt common mechanisms for both NAD+ and nucleic acid substrate binding. The conserved SGR motif induces a significant conformational change in the donor loop upon NAD+ binding to facilitate the catalytic reaction of ART. Moreover, the nucleic acid-binding residue redundancy provides structural flexibility to accommodate different nucleic acid substrates. Mutational assays revealed that TRPT1s employ different catalytic and nucleic acid-binding residues to perform nucleic acid ADP-ribosylation and RNA 2'-phosphotransferase activities. Finally, cellular assays revealed that the mammalian TRPT1 is able to promote endocervical HeLa cell survival and proliferation. Together, our results provide structural and biochemical insights into the molecular mechanism of TRPT1 for nucleic acid ADP-ribosylation.

Discovery of a miniaturized PROTAC with potent activity and high selectivity.

The approved small-molecule inhibitors of anaplastic lymphoma kinase (ALK) have shown remarkable efficacy in some subset of cancer patients. However, the numerous ALK mutants or fusion partners are resistant to such drugs, greatly limiting their application in clinic. Despite the drug design strategy of proteolysis-targeting chimera (PROTAC) holds great potential to overcome drug resistance in theory, there are obvious disadvantages for the reported PROTACs that include high molecular weight, long linkers, difficult synthesis routes as well as insufficient evidence in activity for diverse ALK mutants. In this study, we designed and synthesized a miniaturized PROTAC of ALK named AP-1 following the principle of minimalist design. Two simple chemical units of ligands and a minimized linker with only two atoms were selected for synthesis of AP-1. At cellular level, AP-1 successfully degraded three types of ALK mutants including NPM-ALK, EML4-ALK and F1174L mutation ALK form with potent activity, high selectivity in ALK-positive cells. In xenograft mouse model, AP-1 showed the stronger antitumor efficacy than ceritinib as well as ALK degraders reported in literatures. AP-1 with an extremely simple PROTAC structure can be served as an effective candidate drug for therapy of various types of ALK-positive cancers. And the design principle of AP-1 has a good guiding significance for overcoming the disadvantages such as excessive molecular weight and poor solubility of PROTAC.

Spatial distribution and ecological risk assessment of soil heavy metals in a typical volcanic area: Influence of parent materials.

To understand the distribution characteristics and potential ecological risks of heavy metals in soils in the typical volcanic area, 2,592 soil samples were collected from the surface layer (0-20 cm) and 269 samples were collected from the middle (80-100 cm) and deep layers (180-200 cm) in northeast of Hainan province, China. Accordingly, eight heavy metals (Cu, Pb, Zn, Cr, Ni, Cd, As, and Hg) were analyzed and determined. The effects of different parent materials and land use types on the accumulation of heavy metals in soils were compared, and the primary heavy metal sources were analyzed. The pollution level and ecological risk of heavy metals in soils in the study area were evaluated using the geo-accumulation index (Igeo) and potential ecological risk indices (E i & RI). The results showed that, except that of Pb, the median concentrations of the analyzed heavy metals in the surface soils were higher than the background concentrations in the Hainan Island soils, indicating varying degrees of accumulation. The influence of land use type on the accumulation of heavy metals in surface soils varied from that of the parent materials. Anthropogenic activities highly influenced As, Cd, Hg, and Pb concentrations, whereas geological conditions primarily influenced Cr, Cu, Ni, and Zn concentrations. The Igeo results showed that the mean value of the eight metal elements were greater than zero, except for Pb. In the surface soils, the Igeo values of As, Cd, Hg, and Zn mostly fell into the light to moderate pollution class, and those of Cr, Cu, and Ni fell into the medium and heavy pollution class. The RI of the study area showed a high to significantly high ecological risk because of the Cd, Hg, and Ni concentrations. The results give a new insight in the parent material's geochemical control on the heavy metal elements in soils, and it can serve as a reference for the background value of local soil heavy metals and provide a scientific basis for controlling the potential ecological risk of heavy metals and reasonable land use plans.

Vertical distribution and major influencing factors of soil selenium in tropical climate: A case study of Chengmai County, Hainan Island.

Soil selenium is of great significance to human health. Soil-forming parent rocks are the most critical factor that influences soil Se levels. Chengmai County, Hainan Island, has a tropical climate and diverse types of parent rocks, in which soil Se content is high. This study investigated the vertical distribution of soil Se from various parent rock substrates under tropical climatic conditions, and the factors that influence these soil Se contents, with 69 vertical soil profiles covering Chengmai County. The vertical distribution of soil Se and correlations with CIA (chemical alteration index), Al2O3, TFe2O3 (total iron oxide expressed as Fe2O3), total iodine, SOC (soil organic carbon), and pH were analysed. As per the results, the mean ± standard error of Se content in the A, B, and C horizons was 0.88 ± 0.13 mg/kg, 0.77 ± 0.08 mg/kg and 0.45 ± 0.05 mg/kg, respectively. The parent rock strictly controlled the horizon distribution of Se in the A-horizon. Soil Se showed A-B-horizons-enrichment in the vertical profile, especially in soil profiles overlying granite and basalt. It is hypothesised that the Se enriched in soils developed from the Tuolie Formation due to the release of Se from the weathering process of Se-rich rocks. Meanwhile, Se in soils developed from granite and basalt is more closely associated with exogenous input. Another crucial factor for the high level of Se in Chengmai County is the tropical climate, which has led the rocks to generally undergo intense chemical weathering. This results in soils rich in clay minerals and Fe/Al oxyhydroxides, which easily absorb and retain Se. Furthermore, the Se content of the B-horizon was generally higher than that of the A-horizon due to leaching. These results provide further knowledge and understanding of the geochemical behaviour of soil Se and guide the evaluation of Se-rich land resources under tropical climatic conditions.

[Enrichment Factors, Health Risk, and Source Identification of Heavy Metals in Agricultural Soils in Semi-arid Region of Hainan Island].

To understand the enrichment factors and pollution levels of heavy metals in agricultural soils in the semi-arid region of Hainan island, 1818 surface soil samples were collected in Gancheng Town and analyzed for their heavy metal contents and physicochemical composition. Correlation analysis was used to determine the heavy metal enrichment factors. The geo-accumulation index (Igeo), comprehensive ecological risk index (RI), and hazard index (HI), as well as carcinogenic risk (CR), were used to assess the degree of pollution and health risk. Positive matrix factorization (PMF) was used to determine the primary sources of pollution and priority sources. The average values of heavy metal contents in the topsoil were 22.7, 0.128, 33.4, 14.5, 0.032, 9.32, 32.5, and 43.3 mg·kg-1 for As, Cd, Cr, Cu, Hg, Ni, Pb, and Zn, respectively. With the exception of Zn, the concentrations of other heavy metals in the topsoil were higher than the soil background values of Hainan, showing different degrees of heavy metal accumulation effect. The Igeo revealed that the major pollutant element in soils was As, followed by Cd and Cu. The RI showed that the proportion of soil samples that were high-risk level or worse was 29.4% of the total number of samples, among which As was the major source of risk. The health risk assessment results indicated that As, Cr, and Ni exposure presented carcinogenic risk for children with high CR values. Based on PMF, four major sources of heavy metals were identified in the study area. Hg was derived mainly from industrial sources, and As was closely associated with agricultural activities. Ni, Cu, Cr, and Zn were related to soil parent materials. Pb and Cd were associated with agricultural activities and traffic emissions. The PMF models combined with correlation analysis were useful for estimating the source apportionment of heavy metals in soils.

TriD-LAMP: A pump-free microfluidic chip for duplex droplet digital loop-mediated isothermal amplification analysis.

Digital nucleic acid amplification techniques are powerful and attractive approaches for providing sensitive and absolute quantification in biology. Among these, digital loop-mediated isothermal amplification (dLAMP) shows the potential for field detection, since its robustness and independence from thermal cycling. The key of dLAMP is to generate a large number of individual droplets or microwells. However, an auxiliary precision pump is always required for sample digitalization. In addition, current systems for droplet dLAMP usually need to transfer the droplets after digitalization or amplification. Herein, we developed and evaluated a pump-free microfluidic chip for duplex droplet dLAMP (TriD-LAMP) detection. This chip was designed based on step emulsification and contains a droplet generation zone and a droplet storage zone. Droplets are formed through the step due to the difference in Laplace pressure. There are 64 parallel nozzles that could generate tens of thousands of uniform droplets manually (variation <5%). The storage zone for droplets collection was previously filled with oil containing fluorosurfactant that keeps the droplets from fusing and evaporation during the amplification. Therefore, this custom chip is able to perform all stages of the dLAMP process without transferring droplets. Combined with the optimized fluorescent probe method, the chip achieves accurate quantification of the E. coli DNA down to 19.8 copies/µL. As a proof of concept, the simultaneous quantification of two targets was successfully realized on this custom chip. Conclusively, this integrated, pump-free TriD-LAMP chip can provide a promising tool for multiple targets detection in clinical diagnostics and point-of-care applications.

A review: research progress on the formation mechanism of porous anodic oxides.

Owing to the great development potential of porous anodic oxides (PAO) in many fields, research on their formation mechanisms, fabrication processes and applications has a history of more than ten years. Although compared with research on the fabrication processes and applications of PAO, research on their formation mechanisms started late, several mainstream theories have been formed in the academic community, including the field-assisted dissolution (FAD) theory, the field-assisted ejection (FAE) theory, the self-organization theory, the ionic and electronic current theory and the oxygen bubble mould effect. This review will focus on summarizing the core views of the mainstream mechanisms mentioned above and comparing the explanations for some of their classical experimental phenomena.