Optical fiber surface plasmon resonance sensor using electroless-plated gold film for thrombin detection.

This paper describes the simple and label-free detection of thrombin using optical fiber surface plasmon resonance (SPR) sensors based on gold films prepared by the cost-effective method of electroless plating. The plating conditions for simultaneously obtaining gold film on cylindrical core and end surfaces of an optical fiber suitable for measurement were optimized. The fabricated sensor exhibited a linear refractive index sensitivity of 2150 nm/RIU and 7.136 (a.u.)/RIU in the refractive index of 1.3329-1.3605 interrogated by resonance wavelength and amplitude methods respectively and a single wavelength monitoring method was proposed to investigate the sensing performance of this sensor. Polyadenine diblock and thiolated thrombin aptamers were immobilized on gold nanoparticles and gold films respectively to implement a sandwich optical fiber assay for thrombin. The developed optical fiber SPR sensors were successfully used in the determination of thrombin down to 0.56 nM over a wide range from 2 to 100 nM and showed good selectivity for thrombin, which indicated their potential clinical applications for biomedical samples.

Association of cardiotrophin-like cytokine factor 1 levels in peripheral blood mononuclear cells with bone mineral density and osteoporosis in postmenopausal women.

BACKGROUND: Recent research has suggested that cardiotrophin-like cytokine factor 1 (CLCF1) may be an important regulator of bone homeostasis. Furthermore, a whole gene chip analysis suggested that the expression levels of CLCF1 in the peripheral blood mononuclear cells (PBMCs) were downregulated in postmenopausal women with osteoporosis. This study aimed to assess whether the expression levels of CLCF1 in PBMCs can reflect the severity of bone mass loss and the related fracture risk. METHODS: In all, 360 postmenopausal women, aged 50 to 80 years, were included in the study. A survey to evaluate the participants' health status, measurement of bone mineral density (BMD), routine blood test, and CLCF1 expression level test were performed. RESULTS: Based on the participants' bone health, 27 (7.5%), 165 (45.83%), and 168 (46.67%) participants were divided into the normal, osteopenia, and osteoporosis groups, respectively. CLCF1 protein levels in the normal and osteopenia groups were higher than those in the osteoporosis group. While the CLCF1 mRNA level was positively associated with the BMD of total femur (r = 0.169, p = 0.011) and lumbar spine (r = 0.176, p = 0.001), the protein level was positively associated with the BMD of the lumbar spine (r = 0.261, p < 0.001), femoral neck (r = 0.236, p = 0.001), greater trochanter (r = 0.228, p = 0.001), and Ward's triangle (r = 0.149, p = 0.036). Both the mRNA and protein levels were negatively associated with osteoporosis development (r = - 0.085, p = 0.011 and r = - 0.173, p = 0.014, respectively). The association between CLCF1 protein level and fracture risk was not significant after adjusting for BMD. CONCLUSIONS: To our knowledge, this is the first clinical study to show that CLCF1 expression levels in the PBMCs of postmenopausal women can reflect the amount of bone mass or the severity of bone mass loss.

Sevoflurane prevents vulnerable plaque disruption in apolipoprotein E-knockout mice by increasing collagen deposition and inhibiting inflammation.

BACKGROUND: Sevoflurane may reduce the occurrence of major adverse cardiovascular events (MACCEs) in surgical patients, although the mechanisms are poorly understood. We hypothesised that sevoflurane stabilises atherosclerotic plaques by inhibiting inflammation and enhancing prolyl-4-hydroxylase α1 (P4Hα1), the rate-limiting subunit for the P4H enzyme essential for collagen synthesis. METHODS: We established a vulnerable arterial plaque model in apolipoprotein E-knockout mice (ApoE-/-) fed a high-fat diet that underwent daily restraint/noise stress, with/without a single prior exposure to sevoflurane for 6 h (1-3%; n=30 per group). In vitro, smooth muscle cells (SMCs) were incubated with tumour necrosis factor-alpha in the presence/absence of sevoflurane. Immunohistochemistry, immunoblots, and mRNA concentrations were used to quantify the effect of sevoflurane on plaque formation, expression of inflammatory cytokines, P4Hα1, and collagen subtypes in atherosclerotic plaques or isolated SMCs. RESULTS: In ApoE-/- mice, inhalation of sevoflurane 1-3% for 6 h reduced the aortic plaque size by 8-29% in a dose-dependent manner, compared with control mice that underwent restraint stress alone (P<0.05); this was associated with reduced macrophage infiltration and lower lipid concentrations in plaques. Sevoflurane reduced gene transcription and protein expression levels of pro-inflammatory cytokines (≥69-75%; P<0.05) and matrix metalloproteinases (≥39-65%; P<0.05) at both gene transcription and protein levels, compared with controls. Sevoflurane dose dependently increased Types I and III collagen deposition through enhanced protein expression of P4Hα1, both in vivo and in vitro (0.7-3.3-fold change; P<0.05). CONCLUSIONS: Sevoflurane dose dependently promotes plaque stabilisation and decreases the incidence of plaque disruption in ApoE-/- mice by increasing collagen deposition and inhibiting inflammation. These mechanisms may contribute to sevoflurane reducing MACCE.

Immune recognition, antimicrobial and opsonic activities mediated by a sialic acid binding lectin from Ruditapes philippinarum.

In the present study, a sialic acid-binding lectin was identified and characterized from Manila clam Ruditapes philippinarum (designed as RpSABL-1). Multiple alignments strongly suggested that RpSABL-1 was a new member of the sialic acid-binding lectin family. In non-stimulated clams, RpSABL-1 transcripts were constitutively expressed in all five tested tissues, especially in hepatopancreas. After Vibrio anguillarum challenge, the expression of RpSABL-1 mRNA was significantly up-regulated at 6 h (P < 0.05), 12 h (P < 0.01) and 24 h (P < 0.01). Recombinant RpSABL-1 protein (rRpSABL-1) displayed apparent binding activities towards lipopolysaccharides (LPS) and peptidoglycan (PGN), but not to glucan or chitin in vitro. Coinciding with the PAMPs binding assay, rRpSABL-1 exhibited obvious agglutination activities against Gram-positive bacterium Staphyloccocus aureus, Gram-negative bacteria Escherichia coli, V. anguillarum and Vibrio harveyi. Meanwhile, rRpSABL-1 showed antibacterial activities against E. coli, and biofilm formation of E. coli could also be inhibited after incubated with rRpSABL-1. Moreover, the encapsulation, phagocytosis and chemotactic ability of hemocytes could be enhanced by rRpSABL-1. All these results suggested that RpSABL-1 could function as a pattern recognition receptor with versatile functions in the innate immune responses of R. philippinarum.

Voltammetric determination of DNA based on regulation of DNA strand displacement using an allosteric DNA toehold.

An electrochemical biosensor for determination of DNA is described that is based on the reaction of regulated DNA (reg-DNA) first with substrated DNA (subs-DNA) to form a reaction intermediate. The intermediate binds target DNA (T) by hybridization and initiates a branch migration leading to the production of complex of substrated DNA and target DNA (TC). Once TC is produced, it reacts with assisted DNA (ass-DNA) through a toehold exchange mechanism, yielding the product complex of substrated DNA and assisted DNA (CS). The target is then released back into the solution and and catalyzes the next cycle of toehold-exchange with the reaction intermediate of substrated DNA and regulated DNA (CPR). Unlike in a conventional DNA toehold that is hardwired with the branch migration domain, the allosteric DNA toehold is designed into a reg-DNA which is independent of the branch migration domain. Under the optimal experimental conditions and at a working potential as low as 0.18 V, response to DNA is linear in the 1 fM to 1000 pM concentration range, and the detection limit is 0.83 fM. The assay is highly specific and can discriminate target DNA even from a single-base mismatch. It was applied to the analysis of DNA spiked plasma samples. Graphical abstract Schematic illustration of the electrochemical strategy for target DNA detection based on regulation of DNA strand displacement using an allosteric DNA toehold strategy. It can be used to analyze DNA-spiked plasma samples and has a low detection limit of 0.83 fM.

Optimal design of depth-scaling error for multilayer diffractive optical elements with oblique incidence.

The mathematical analytic models of the relationship between depth-scaling error and diffraction efficiency/polychromatic integral diffraction efficiency (PIDE) for multilayer diffractive optical elements (MLDOEs) with oblique incidence are presented. A method for optimal design of the depth-scaling error for MLDOEs with consideration of comprehensive PIDE working within a range of incident angle is established. The effect of depth-scaling error on diffraction efficiency and PIDE of MLDOEs working in the visible wavelength with oblique incidence is analyzed. For the MLDOEs working within the range of incident angle 0-20°, the maximum comprehensive PIDE is 98.24%, and the optimal relative depth-scaling error is -6.55%. The analytic method and conclusion provide a theoretical basis for the determination of manufacturing tolerance of depth-scaling error for MLDOEs with oblique incidence.

Ggnbp2 Is Essential for Pregnancy Success via Regulation of Mouse Trophoblast Stem Cell Proliferation and Differentiation.

The Ggnbp2 null mutant embryos died in utero between Embryonic Days 13.5 to 15.5 with dysmorphic placentae, characterized by excessive nonvascular cell nests consisting of proliferative trophoblastic tissue and abundant trophoblast stem cells (TSCs) in the labyrinth. Lethality of Ggnbp2 null embryos was caused by insufficient placental perfusion as a result of remarkable decreases in both fetal and maternal blood vessels in the labyrinth. These defects were accompanied by a significant elevation of c-Met expression and phosphorylation and its downstream effector Stat3 activation. Knockdown of Ggnbp2 in wild-type TSCs in vitro provoked the proliferation but delayed the differentiation with an upregulation of c-Met expression and an enhanced phosphorylation of c-Met and Stat3. In contrast, overexpression of Ggnbp2 in wild-type TSCs exhibited completely opposite effects compared to knockdown TSCs. These results suggest that loss of GGNBP2 in the placenta aberrantly overactivates c-Met-Stat3 signaling, alters TSC proliferation and differentiation, and ultimately compromises the structure of placental vascular labyrinth. Our studies for the first time demonstrate that GGNBP2 is an essential factor for pregnancy success acting through the maintenance of a balance of TSC proliferation and differentiation during placental development.

Angiotensin 1-7 restrains vascular injury of extracorporeal membrane oxygenation by inhibiting ferroptosis.

BACKGROUND: Angiotensin 1-7 (Ang1-7) is the classical end product of angiotensin II, which has the effects of dilating blood vessels, protecting endothelial cells, anti-hypertension, improving cardiac function, and inhibiting atherosclerosis. We hypothesize that Ang1-7 inhibits human umbilical vein endothelial cells (HUVEC) ferroptosis through NF-κB/P53 signal pathway, and reduces extracorporeal membrane oxygenation (ECMO) vascular injury. METHODS: Cultured HUVEC were seeded into 15 wells and randomly divided into five groups: the control group and four experimental groups (erastin, erastin + Ang1-7, erastin + Ang1-7 + Betulinic acid, erastin + Betulinic acid). After stimulation, cell viability, lactate dehydrogenase (LDH), malondialdehyde (MDA), and superoxide dismutase (SOD) activity were measured. The effects of Ang1-7 on HUVEC microstructure, antioxidant enzymes (ferritin heavy chain 1 (FTH1), cystine/glutamic acid reverse transport solute carrier family 7 members 11 (SLC7A11 or XCT), superoxide dismutase-2 (SOD-2) and glutathione peroxidase 4 (GPX4)), NF-κB, P-NF-κB, P53, and P-P53). RESULTS: Erastin stimulation promoted HUVEC lipid peroxidation, decreased antioxidant enzyme expression, increased P-NF-κB, P53, and P-P53 expressions, and damaged HUVEC mitochondrial structure. Ang1-7 alleviated the effect of erastin on HUVEC, which was destroyed by Betulinic acid. CONCLUSION: Angiotensin1-7 pretreatment inhibited vascular endothelial cells' ferroptosis and alleviated ECMO vessel injury through NF-κB /P53 signal pathway.

Cope with copper: From molecular mechanisms of cuproptosis to copper-related kidney diseases.

Cuproptosis has recently been identified as a novel regulatory mechanism of cell death. It is characterized by the accumulation of copper in mitochondria and its binding to acylated proteins. These characteristics lead to the downregulation of iron-sulfur cluster proteins and protein toxicity stress, ultimately resulting in cell death. Cuproptosis is distinct from other types of cell death, including necrosis, apoptosis, ferroptosis, and pyroptosis. Cu induces oxidative stress damage, protein acylation, and the oligomerization of acylated TCA cycle proteins. These processes lead to the downregulation of iron-sulfur cluster proteins and protein toxicity stress, disrupting cellular Cu homeostasis, and causing cell death. Cuproptosis plays a significant role in the development and progression of various kidney diseases such as acute kidney injury, chronic kidney disease, diabetic nephropathy, kidney transplantation, and kidney stones. On the one hand, inducers of cuproptosis, such as disulfiram (DSF), chloroquinolone, and elesclomol facilitate cuproptosis by promoting cell oxidative stress. In contrast, inhibitors of Cu chelators, such as tetraethylenepentamine and tetrathiomolybdate, relieve these diseases by inhibiting apoptosis. To summarize, cuproptosis plays a significant role in the pathogenesis of kidney disease. This review comprehensively discusses the molecular mechanisms underlying cuproptosis and its significance in kidney diseases.

Comparisons of gene expression between peripheral blood mononuclear cells and bone tissue in osteoporosis.

Osteoporosis (OP) is one of the major public health problems in the world. However, the biomarkers between the peripheral blood mononuclear cells (PBMs) and bone tissue for prognosis of OP have not been well characterized. This study aimed to explore the similarities and differences of the gene expression profiles between the PBMs and bone tissue and identify potential genes, transcription factors (TFs) and hub proteins involved in OP. The patients were enrolled as an experimental group, and healthy subjects served as normal controls. Human whole-genome expression chips were used to analyze gene expression profiles from PBMs and bone tissue. And the differentially expressed genes (DEGs) were subsequently studied using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis. The above DEGs were constructed into protein-protein interaction network. Finally, TF-DEGs regulation networks were constructed. Microarray analysis revealed that 226 DEGs were identified between OP and normal controls in the PBMs, while 2295 DEGs were identified in the bone tissue. And 13 common DEGs were obtained by comparing the 2 tissues. The Gene Ontology analysis indicated that DEGs in the PBMs were more involved in immune response, while DEGs in bone were more involved in renal response and urea transmembrane transport. And the Kyoto Encyclopedia of Genes and Genomes analysis indicated almost all of the pathways in the PBMs were overlapped with those in the bone tissue. Furthermore, protein-protein interaction network presented 6 hub proteins: PI3K1, APP, GNB5, FPR2, GNG13, and PLCG1. APP has been found to be associated with OP. Finally, 5 key TFs were identified by TF-DEGs regulation networks analysis (CREB1, RUNX1, STAT3, CREBBP, and GLI1) and were supposed to be associated with OP. This study enhanced our understanding of the pathogenesis of OP. PI3K1, GNB5, FPR2, GNG13, and PLCG1 might be the potential targets of OP.

The molecular mechanisms of cuproptosis and its relevance to cardiovascular disease.

Recently, cuproptosis has been demonstrated to be a new non-apototic cell death mode that is characterized by copper dependence and the regulation of mitochondrial respiration. Cuproptosis is distinct from known cell death modes such as apoptosis, necrosis, pyroptosis, or ferroptosis. Excessive copper induces cuproptosis by promoting protein toxic stress reactions via copper-dependent anomalous oligomerization of lipoylation proteins in the tricarboxylic acid (TCA) cycle and reducing iron-sulfur cluster protein levels. Ferredoxin1 (FDX1) promotes dihydrolipoyl transacetylase (DLAT) lipoacylation and abates iron-sulfur cluster proteins by reducing Cu2+ to Cu+, inducing cell death. Copper homeostasis depends on the copper transporter, and disturbances to this homeostasis cause cuproptosis. Recent evidence has shown that cuproptosis plays a significant role in the occurrence and development of many cardiovascular diseases, such as myocardial ischemia/reperfusion (I/R) injury, heart failure, atherosclerosis, and arrhythmias. Copper chelators, such as ammonium tetrathiomolybdate(VI) and DL-Penicillamine, may ease the above cardiovascular diseases by inhibiting the cuproptosis pathway. Oxidative phosphorylation inhibitors may inhibit cuproptosis by inhibiting protein stress response. In conclusion, cuproptosis plays an essential role in cardiovascular disease pathogenesis. Inhibition of cardiovascular cuproptosis is expected to become a potential treatment. Here, we will thoroughly review the molecular mechanisms involved in cuproptosis and its significance in cardiovascular disease.

Establishment of a Real-Time Recombinase Polymerase Amplification for Rapid Detection of Pathogenic Yersinia enterocolitica.

Yersinia enterocolitica is a zoonotic proto-microbe that is widespread throughout the world, causes self-limiting diseases in humans or animals and even leads to sepsis and death in patients with severe cases. In this study, a real-time recombinase polymerase amplification (RPA) assay for pathogenic Y. enterocolitica was established based on the ail gene. The results showed that the RPA detection for Y. enterocolitica could be completed within 20 min at an isothermal temperature of 38 °C by optimizing the conditions in the primers and Exo probe. Moreover, the sensitivity of the current RT-RPA was 10-4 ng/µL, and the study found that the assay was negative in the application of the genomic DNA of other pathogens. These suggest the establishment of a rapid and sensitive real-time RPA method for the detection of pathogenic Y. enterocolitica, which can provide new understandings for the early diagnosis of the pathogens.

Integrated proteomics and metabolomics analysis of lumbar in a rat model of osteoporosis treated with Gushukang capsules.

BACKGROUND: Gushukang (GSK) capsules are a Chinese patented medicine that is widely used in clinics for the treatment of osteoporosis (OP). Animal experiments have revealed that the bone mineral density of osteoporotic rats increase after treatment with GSK capsules. However, the specific mechanism and target of GSK in the treatment of osteoporosis are unclear. Further studies are needed. METHODS: Metabolomics (GC/MS) and proteomics (TMT-LC-MC/MC) with bioinformatics (KEGG pathway enrichment), correlation analysis (Pearson correlation matrix), and joint pathway analysis (MetaboAnalyst) were employed to determine the underlying mechanisms of GSK. The differential expression proteins were verified by WB experiment. RESULTS: The regulation of proteins, i.e., Cant1, Gstz1, Aldh3b1, Bid, and Slc1a3, in the common metabolic pathway of differential proteins and metabolites between GSK/OP and OP/SHAM was corrected in the GSK group. The regulation of 12 metabolites (tyramine, thymidine, deoxycytidine, cytosine, L-Aspartate, etc.) were differential in the common enrichment metabolic pathway between GSK /OP and OP/SHAM. Differential proteins and metabolites jointly regulate 11 metabolic pathways, such as purine metabolism, pyrimidine metabolism, histidine metabolism, beta-alanine metabolism, and so on. CONCLUSION: GSK may protect bone metabolism in osteoporotic rats by affecting nucleotide metabolism, amino acid metabolism, and the immune system.

Corrigendum: Propofol Protects Myocardium From Ischemia/Reperfusion Injury by Inhibiting Ferroptosis Through the AKT/p53 Signaling Pathway.

[This corrects the article DOI: 10.3389/fphar.2022.841410.].

Propofol Protects Myocardium From Ischemia/Reperfusion Injury by Inhibiting Ferroptosis Through the AKT/p53 Signaling Pathway.

The molecular mechanism underlying the protective role of propofol against myocardial ischemia/reperfusion (I/R) injury remains poorly understood. Previous studies have shown that ferroptosis is an imperative pathological process in myocardial I/R injury. We hypothesized that propofol prevents myocardial I/R injury by inhibiting ferroptosis via the AKT/p53 signaling pathway. The ferroptosis-inducing agent erastin (E) and AKT inhibitor MK2206 (MK) were used to investigate the role of propofol in myocardial I/R injury. H9C2 cells treated without any reagents, erastin for 24 h, propofol for 1 h before adding erastin were assigned as the control (C), E, and E + P group, respectively. Cell viability, reactive oxygen species (ROS), and the expression of antioxidant enzymes, including ferritin heavy chain 1 (FTH1), cysteine/glutamate transporter (XCT), and glutathione peroxidase 4 (GPX4) in H9C2 cells. Rat hearts from the I/R + P or I/R groups were treated with or without propofol for 20 min before stopping perfusion for 30 min and reperfusion for 60 min. Rat hearts from the I/R + P + MK or I/R + MK groups were treated with or without propofol for 20 min, with a 10-min treatment of MK2206 before stopping perfusion. Myocardial histopathology, mitochondrial structure, iron levels, and antioxidant enzymes expression were assessed. Our results demonstrated that erastin increased H9C2 cell mortality and reduced the expression of antioxidant enzymes. I/R, which reduced the expression of antioxidant enzymes and increased iron or p53 (p < 0.05), boosted myocardium pathological and mitochondrion damage. Propofol inhibited these changes; however, the effects of propofol on I/R injury were antagonized by MK (p < 0.05). In addition, AKT siRNA inhibited the propofol-induced expression of antioxidant enzymes (p < 0.05). Our findings confirm that propofol protects myocardium from I/R injury by inhibiting ferroptosis via the AKT/p53 signal pathway.

Dysregulation of Notch-FGF signaling axis in germ cells results in cystic dilation of the rete testis in mice.

Numb (Nb) and Numb-like (Nbl) are functionally redundant adaptor proteins that critically regulate cell fate and morphogenesis in a variety of organs. We selectively deleted Nb and Nbl in testicular germ cells by breeding Nb/Nbl floxed mice with a transgenic mouse line Tex101-Cre. The mutant mice developed unilateral or bilateral cystic dilation in the rete testis (RT). Dye trace indicated partial blockages in the testicular hilum. Morphological and immunohistochemical evaluations revealed that the lining epithelium of the cysts possessed similar characteristics of RT epithelium, suggesting that the cyst originated from dilation of the RT lumen. Spermatogenesis and the efferent ducts were unaffected. In comparisons of isolated germ cells from mutants to control mice, the Notch activity considerably increased and the expression of Notch target gene Hey1 significantly elevated. Further studies identified that germ cell Fgf4 expression negatively correlated the Notch activity and demonstrated that blockade of FGF receptors mediated FGF4 signaling induced enlargement of the RT lumen in vitro. The crucial role of the FGF4 signaling in modulation of RT development was verified by the selective germ cell Fgf4 ablation, which displayed a phenotype similar to that of germ cell Nb/Nbl null mutant males. These findings indicate that aberrant over-activation of the Notch signaling in germ cells due to Nb/Nbl abrogation impairs the RT development, which is through the suppressing germ cell Fgf4 expression. The present study uncovers the presence of a lumicrine signal pathway in which secreted/diffusible protein FGF4 produced by germ cells is essential for normal RT development.

Experimental demonstration of a controllable electrostatic molecular beam splitter.

We experimentally demonstrate a controllable electrostatic beam splitter for guided ND3 molecules with a single Y-shaped charged wire and a homogeneous bias field generated by a charged metallic parallel-plate capacitor. We study the dependences of the splitting ratio R of the guided ND3 beam and its relative guiding efficiency η on the voltage difference between two output arms of the splitter. The influences of the molecular velocity v and the cutting position L on the splitting ratio R are investigated as well, and the guiding and splitting dynamic processes of cold molecules are simulated. Our study shows that the splitting ratio R of our splitter can be conveniently adjusted from 10% to 90% by changing ΔU from -6 kV to +6 kV, and the simulated results are consistent with our experimental ones.

Law of coal caving behind the flexible shield support in pseudo-inclined working face.

Complex boundary conditions are the major influencing factors of coal caving law in the pseudo-inclined working face. The main purpose of this study is to analyze coal caving law of flexible shield support and then to establish the internal relations among coal caving parameters under complex boundary conditions. Firstly, the law of coal caving in different falling modes is simulated physically. Secondly, the coal caving shape, displacement field, and contact force field is simulated. Then, coal caving law and process parameters is analyzed theoretically. Finally, the test was performed in Bai-Ji Mine. The research shows that ellipsoidal ore drawing theory has universal applicability in coal drawing law analysis and parameter optimization. After the Isolated Extraction Zone and Isolated Movement Zone reach the roof, the expansion speed is marked by a short delay, and then, while expanding to the floor, two butted incomplete ellipsoids are formed. There is a time-space difference in coal caving after the support, and some coal will be mined in the next round of coal caving. There are obvious differences in the coal loosening range, displacement field, and contact force field on both sides of the long axis. When the support falls along with the bottom plate, it is more conducive to the release of coal. The test shows that the research is of great significance for optimizing the caving parameters of flexible shield support in the pseudo-inclined working face of the steep seam.

An ultrasensitive and specific electrochemical biosensor for DNA detection based on T7 exonuclease-assisted regulatory strand displacement amplification.

An electrochemical biosensor for determination of DNA is developed based on T7 exonuclease-assisted regulatory strand displacement dual recycling signal amplification strategy. First, the hairpin probe recognized and bound the target DNA to form a double strand nucleotide structure, and then the T7 exonuclease was introduced. After be digested by T7 exonuclease, the target DNA was released and entered the next cycle of T7 exonuclease-assisted recycle amplification, while accompanied by a large number of mimic targets (output DNAs) into another cycle. Second, the mimic target reacted with double-chain substrated DNA (CP) by a regulated toehold exchange mechanism, yielding the product complex of detection probes with the help of assisted DNA (S). Finally, after many cycles, a large number of detection probes were produced for binding numerous streptavidin-alkaline phosphatases. The electrochemical biosensor showed very high sensitivity and selectivity with a dynamic response ranged from 0.1 fM to 10 pM with a detection limit of 31.6 aM. Furthermore, this proposed biosensor was successfully applied to the detection of target DNA in 20% diluted serum. The developed strategy has been demonstrated to have the potential for application in molecular diagnostics.

Fluorescence sensing strategy based on aptamer recognition and mismatched catalytic hairpin assembly for highly sensitive detection of alpha-fetoprotein.

At present, alpha fetoprotein (AFP) is mainly used as a serum marker of primary Hepatocellular carcinoma. A simple, enzyme-free sensing strategy is introduced for highly sensitive fluorescence detection of AFP. This detection strategy is based on aptamer recognition and mismatched catalytic hairpin assembly (MCHA). At first, Trigger is locked by aptamer before the introduction of AFP in this aptamer-MCHA system. The aptamer preferentially combines with AFP via powerful attraction in the presence of AFP. This results in the release of trigger and initiation of MCHA cycle, thus forming the H1 and H2 double chain complexes (denoted as H1@H2). Finally, H1@H2 and double chain structure containing fluorophore and its quenched group- BHQ1 (denoted as F@Q) initiated displacement reaction, which caused double chain separation and fluorescence recovery. This assay produces a wide detection range, which is from 0.1 ng mL-1 to 10 µg mL-1 and the limit of detection as 0.033 ng mL-1. The whole detection process was performed at 37 °C for 60 min. In addition, this assay had high anti-interference ability and could be used to detect AFP in clinical serum. This novel AFP detection strategy is able to screen of Hepatocellular carcinoma.