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1.
Mikrochim Acta ; 191(6): 330, 2024 05 14.
Article in English | MEDLINE | ID: mdl-38744738

ABSTRACT

In view of a large number of people infected with Helicobacter pylori (H. pylori) with great harm followed, there is an urgent need to develop a non-invasive, easy-to-operate, and rapid detection method, and to identify effective sterilization strategies. In this study, highly specific nanoprobes with nanozyme activity, Ag@Pt nanoparticles (NPs) with the antibody, were utilized as a novel lateral flow immunoassay (LFIA). The optical label (Ag@Pt NPs) was enhanced by the introduction of the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) and compared with a gold nanoparticles (Au NPs) optical label. Under the optimal condition, Ag@Pt-LFIA and TMB-enhanced Ag@Pt-LFIA for H. pylori were successfully established, two of which were over twofold and 100-fold more sensitive than conventional visual Au NP-based LFIA, respectively. Furthermore, Ag@Pt NPs with the antibody irradiated with NIR laser (808 nm) at a power intensity of 550 mW/cm2 for 5 min exhibited a remarkable antibacterial effect. The nanoprobes could close to bacteria through effective interactions between antibodies and bacteria, thereby benefiting photothermal sterilization. Overall, Ag@Pt NPs provide promising applications in pathogen detection and therapeutic applications.


Subject(s)
Alloys , Helicobacter pylori , Metal Nanoparticles , Platinum , Silver , Helicobacter pylori/radiation effects , Helicobacter pylori/drug effects , Silver/chemistry , Metal Nanoparticles/chemistry , Platinum/chemistry , Alloys/chemistry , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Immunoassay/methods , Benzidines/chemistry , Gold/chemistry , Humans , Sterilization/methods , Limit of Detection
2.
Anal Chem ; 95(14): 6038-6045, 2023 04 11.
Article in English | MEDLINE | ID: mdl-36972550

ABSTRACT

The soluble epoxide hydrolase (sEH) is possibly both a marker for and target of numerous diseases. Herein, we describe a homogeneous mix-and-read assay for the detection of human sEH based on using split-luciferase detection coupled with anti-sEH nanobodies. Selective anti-sEH nanobodies were individually fused with NanoLuc Binary Technology (NanoBiT), which consists of a large and small portion of NanoLuc (LgBiT and SmBiT, respectively). Different orientations of the LgBiT and SmBiT-nanobody fusions were expressed and investigated for their ability to reform the active NanoLuc in the presence of the sEH. After optimization, the linear range of the assay could reach 3 orders of magnitude with a limit of detection (LOD) of 1.4 ng/mL. The assay has a high sensitivity to human sEH and reached a similar detection limit to our previously reported conventional nanobody-based ELISA. The procedure of the assay was faster (30 min total) and easy to operate, providing a more flexible and simple way to monitor human sEH levels in biological samples. In general, the immunoassay proposed here offers a more efficient detection and quantification approach that can be easily adapted to numerous macromolecules.


Subject(s)
Single-Domain Antibodies , Luciferases/analysis , Humans , Epoxide Hydrolases/metabolism , Time Factors , Solubility , Single-Domain Antibodies/immunology , Calibration , Animals , Mice , Rats
3.
Anal Bioanal Chem ; 414(23): 6939-6946, 2022 Sep.
Article in English | MEDLINE | ID: mdl-35945290

ABSTRACT

Ferritin, widely present in liver and spleen tissue, is considered as a serological biomarker for liver diseases and cancers. The detection of ferritin may be an important tool in health diagnosis. In this study, 14 non-immunized chicken spleens were utilized to construct a single-chain fragment (scFv) phage library. After 4 rounds of panning, 7 unique clones were obtained. The optimal clone was further screened and combined with NanoLuc luciferase (Nluc) as a dual functional immunoprobe to bioluminescent enzyme immunoassay (BLEIA), which was twice as sensitive as its parental scFv-based double-sandwich enzyme-linked immunoassay (ds-ELISA). The cross-reactivity analysis revealed that the proposed methods were highly selective and suitable for clinical detection. To further verify the performance of the immunoassays, serum samples were tested by the proposed methods and a commercial ELISA kit, and there was a good correlation between the results. These results suggested that scFv fused with Nluc might be a powerful dual functional tool for rapid, practically reliable, and highly sensitive ferritin detection.


Subject(s)
Single-Chain Antibodies , Enzyme-Linked Immunosorbent Assay , Ferritins , Immunoassay , Immunoenzyme Techniques , Luciferases/genetics , Peptide Library
4.
Biochem Biophys Res Commun ; 566: 16-23, 2021 08 20.
Article in English | MEDLINE | ID: mdl-34111667

ABSTRACT

With the aggravation of global aging, the rapid rise in the obesity rate, and the increasing number of patients with intervertebral disc degeneration (IDD), the principles and mechanism of this disease remain unclear. This study explored the molecular mechanism of IDD treatment through interactions of the lncRNA-miRNA-mRNA-signaling pathways and the effects on the proliferation and apoptosis of human nucleus pulposus cells (HNPCs) cultured in vitro. Our study revealed that lncRNA JPX is expressed at low levels in HNPCs under normoxic conditions. Luciferase and RNA pull-down assays were used to verify that lncRNA JPX directly bound to miR-18a-5p and influenced HNPC proliferation and apoptosis. Subsequently, a luciferase assay confirmed the direct binding of miR-18a-5p to HIF-1α and demonstrated a negative correlation between miR-18a-5p and HIF-1α. In addition, the HIF-1α antagonist reversed the inhibition of the Hippo-YAP pathway by the miR-18a-5p inhibitor. In conclusion, overexpression of lncRNA JPX upregulated HIF-1α by inhibiting the expression of miR-18a-5p, thereby inhibiting the Hippo-YAP pathway. By inhibiting this pathway, JPX overexpression promoted the proliferation of HNPCs and decreased their apoptosis. Therefore, the lncRNA JPX is a potential new target.


Subject(s)
Apoptosis , Nucleus Pulposus/cytology , RNA, Long Noncoding/genetics , Signal Transduction , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Cell Line , Cell Proliferation , Hippo Signaling Pathway , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , MicroRNAs/genetics , MicroRNAs/metabolism , Nucleus Pulposus/metabolism , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , RNA, Long Noncoding/metabolism , Transcription Factors/genetics , Transcription Factors/metabolism , YAP-Signaling Proteins
6.
Acta Radiol ; 57(12): 1515-1523, 2016 Dec.
Article in English | MEDLINE | ID: mdl-26853685

ABSTRACT

Background To extend the time window for thrombolysis, reducing the time for diagnosis and detection of acute cerebral infarction seems to be warranted. Purpose To evaluate the feasibility of implementing an array spatial sensitivity technique (ASSET)-echo-planar imaging (EPI)-fluid attenuated inversion recovery (FLAIR) (AE-FLAIR) sequence into an acute cerebral infarction magnetic resonance (MR) evaluation protocol, and to assess the diagnostic value of AE-FLAIR combined with three-dimensional time-of-flight MR angiography (3D TOF MRA). Material and Methods A total of 100 patients (68 men, 32 women; age range, 44-82 years) with acute cerebral infarction, including 50 consecutive uncooperative and 50 cooperative patients, were evaluated with T1-weighted (T1W) imaging, T2-weighted (T2W) imaging, FLAIR, diffusion-weighted imaging (DWI), 3D TOF, EPI-FLAIR, and AE-FLAIR. Conventional FLAIR, EPI-FLAIR, and AE-FLAIR were assessed by two observers independently for image quality. The optimized group (AE-FLAIR and 3D TOF) and the control group (T1W imaging, T2W imaging, conventional FLAIR, DWI, and 3D TOF) were compared for evaluation time and diagnostic accuracy. Results One hundred and twenty-five lesions were detected and images having adequate diagnostic image quality were in 73% of conventional FLAIR, 62% of EPI-FLAIR, and 89% of AE-FLAIR. The detection time was 12 ± 1 min with 76% accuracy and 4 ± 0.5 min with 100% accuracy in the control and the optimized groups, respectively. Inter-observer agreements of κ = 0.78 and κ = 0.81 were for the optimized group and control group, respectively. Conclusion With reduced acquisition time and better image quality, AE-FLAIR combined with 3D TOF may be used as a rapid diagnosis tool in patients with acute cerebral infarction, especially in uncooperative patients.


Subject(s)
Cerebral Infarction/diagnostic imaging , Imaging, Three-Dimensional/methods , Magnetic Resonance Imaging/methods , Acute Disease , Adult , Aged , Aged, 80 and over , Brain/diagnostic imaging , Echo-Planar Imaging/methods , Feasibility Studies , Female , Humans , Image Processing, Computer-Assisted/methods , Magnetic Resonance Angiography/methods , Male , Middle Aged , Reproducibility of Results , Sensitivity and Specificity
7.
Medicine (Baltimore) ; 103(27): e38362, 2024 Jul 05.
Article in English | MEDLINE | ID: mdl-38968481

ABSTRACT

Laryngeal carcinoma (LC) is reported to have a higher incidence rate among all types of head and neck cancers around the globe. Mechanisms resulting in the pathogenesis of LC are complicated due to involvement of invasion and metastasis and there is a need to understand this complicated multistep process. Numerous molecules including matrix metalloproteinases (MMPs) are involved in regulating metastatic mechanisms. Furthermore, activation and expression of different classes of MMPs have been observed in multiple pathological and physiological events including inflammation, invasion, and metastasis. Among all members of MMPs, matrix metalloproteinases-2 (MMP-2), and matrix metalloproteinases-9 (MMP-9) have been frequently reported to correlate with tumor pathogenesis. The present study is designed to check the involvement of MMP-2 and MMP-9 in LC pathogenesis. 184 laryngeal tumor samples along with adjacent uninvolved healthy sections were collected to check the expression deregulation of the above-mentioned gene in LC using real-time PCR and immunohistochemistry (IHC). Real-time PCR and IHC analyses showed the significant upregulation of MMP-2 (P < .0001) and MMP-9 (P < .0001) genes in laryngeal tumors compared to controls. Spearman correlation showed the positive correlation of expression deregulation of selected MMPs with advanced TNM stage [MMP-2, (P < .0001); MMP-9, P < .0001] and smoking status [MMP-2 (P < .0001); MMP-9 P < .0001] in laryngeal pathogenesis. Receiver operating curve (ROC) analysis showed the good diagnostic/prognostic value of said markers in laryngeal cancer patients. The present study showed that significant upregulation of selected MMPs was found associated with an increased risk of laryngeal cancer and can act as good diagnostic markers for the detection of said disease.


Subject(s)
Laryngeal Neoplasms , Matrix Metalloproteinase 2 , Matrix Metalloproteinase 9 , Humans , Laryngeal Neoplasms/pathology , Laryngeal Neoplasms/genetics , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 2/metabolism , Matrix Metalloproteinase 9/metabolism , Matrix Metalloproteinase 9/genetics , Retrospective Studies , Male , Middle Aged , Female , Aged , Adult , Biomarkers, Tumor/metabolism , Biomarkers, Tumor/genetics , Neoplasm Staging , Immunohistochemistry , Real-Time Polymerase Chain Reaction , Gene Expression Regulation, Neoplastic , Up-Regulation
8.
Anal Methods ; 16(13): 1862-1869, 2024 03 28.
Article in English | MEDLINE | ID: mdl-38463013

ABSTRACT

Cervical cancer (CC) remains one of the most severe global health challenges affecting women, primarily due to persistent infection with high-risk human papillomavirus (HPV) subtypes, particularly with HPV16 and HPV 18. Effective detection of these high-risk HPV strains is crucial for CC prevention. Current screening programs for HPV DNA include PCR and in situ hybridization, which are accurate and sensitive. However, these approaches demand a high level of expertise, along with expensive instruments and consumables, thus hindering their widespread use. Therefore, there is a compelling demand to develop an efficient, straightforward, and cost-effective method. Herein, we propose a lateral flow immunoassay (LFIA) method based on Au@PdPt nanoparticles for the simultaneous detection and genotyping of HPV16 and HPV18 within 15 min. This innovative approach allows for qualitative assessment by the naked eye and enables semi-quantitative detection through a smartphone. In this study, under optimal conditions, the qualitative visual limits of detection (vLOD) for HPV16 and HPV18 reached 0.007 nM and 0.01 nM, respectively, which were 32-fold and 20-fold more sensitive than conventional AuNPs-LFIA for HPV16 and HPV18, respectively. Meanwhile, semi-quantitative limits of detection (qLOD) for HPV16 and HPV18 were 0.05 nM and 0.02 nM, respectively. In conclusion, our formulated approach represents a significant step forward in HPV detection and genotyping, with the potential to enhance accessibility and effectiveness in the early diagnosis of CC at the point of care and beyond.


Subject(s)
Metal Nanoparticles , Papillomavirus Infections , Uterine Cervical Neoplasms , Female , Humans , Human papillomavirus 18/genetics , Human papillomavirus 16/genetics , Papillomavirus Infections/diagnosis , Papillomavirus Infections/prevention & control , Gold , Uterine Cervical Neoplasms/diagnosis , Uterine Cervical Neoplasms/prevention & control , DNA, Viral/genetics , DNA, Viral/analysis , Immunoassay
9.
Front Nutr ; 11: 1456730, 2024.
Article in English | MEDLINE | ID: mdl-39224187

ABSTRACT

Polyphenols, as subordinate metabolites of plants, have demonstrated significant antibacterial, anti-inflammatory, and antioxidant action in scientific learn. These compounds exert their effects through various mechanisms, containing interference with microbial cell structures, rule of host immune responses, and neutralization of free radicals. This multifaceted activity positions polyphenols as promising candidates for maintaining human health and treating related diseases. Notably, in the context of escalating antibiotic resistance, the antibacterial properties of polyphenols offer innovative avenues for the development of new therapeutic agents. Additionally, their anti-inflammatory and antioxidant effects hold substantial potential for treating inflammatory diseases and mitigating the aging process. This review aims to summarize the latest findings on the biological activities of polyphenols, highlighting their mechanisms of action and potential applications in health and disease management. Furthermore, optimizing polyphenol extraction methods aligns with the goals of sustainable and green processing, reducing environmental impact while enhancing food safety and extending shelf life. Employing advanced analytical techniques, such as spectroscopy and chromatography, can ensure the accurate evaluation of polyphenol content and efficacy. These efforts collectively contribute to the ongoing improvement of food processing practices and product quality, promoting a healthier and more sustainable future in the food industry.

10.
J Colloid Interface Sci ; 673: 893-900, 2024 Nov.
Article in English | MEDLINE | ID: mdl-38908288

ABSTRACT

Cervical cancer is one of the most common gynecological malignancies, with the vast majority of which being caused by persistent infection with Human Papillomavirus (HPV) 16 and 18. The current available HPV detection methods are sensitive and genotyped but are restricted by expensive instruments and skilled personnel. The development of an easy-to-use, rapid, and cost-friendly analysis method for HPV is of great need. Herein, hollow palladium-ruthenium nanocages modified with two oligonucleotides (PdRu capture probes) were constructed for genotyping and simultaneous detection of target nucleic acids HPV16 and HPV18 by dual lateral flow assay (DLFA). PdRu capture probes were endowed with bi-functions for the first time, which could be used to output signals and hybridize target nucleic acids. Under optimized conditions, the PdRu based-DLFA with detection limits of 0.93 nM and 0.19 nM, respectively, exhibited convenient operation, and high sensitivity. Meanwhile, the DLFA achieved excellent rapid detection within 20 min, which was attributed to capture probes that can be directly bound to amplification-free target nucleic acids. Therefore, the development of PdRu-based DLFA can be utilized for rapid, sensitive, and simultaneous genotyping detection of HPV16 and HPV18, showing great application for nucleic acid detection.


Subject(s)
Human papillomavirus 16 , Human papillomavirus 18 , Palladium , Palladium/chemistry , Humans , Human papillomavirus 16/genetics , Human papillomavirus 16/isolation & purification , Human papillomavirus 18/genetics , Human papillomavirus 18/isolation & purification , Ruthenium/chemistry , Nanostructures/chemistry , DNA, Viral/analysis , DNA, Viral/genetics , Surface Properties , Papillomavirus Infections/diagnosis , Papillomavirus Infections/virology , Limit of Detection , Particle Size , Nucleic Acid Hybridization , Human Papillomavirus Viruses
11.
Anal Methods ; 16(10): 1508-1514, 2024 03 07.
Article in English | MEDLINE | ID: mdl-38372146

ABSTRACT

Hepatocellular carcinoma (HCC) remains a leading cause of cancer-related mortality globally, ranking third in cancer deaths. Early diagnosis of HCC markers is imperative for effective prognosis and treatment. This study explores the utility of glycocholic acid (GCA) and alpha-fetoprotein (AFP) as biomarkers for liver diseases, with a specific focus on their simultaneous detection for enhanced diagnostic and prognostic capabilities. Harnessing the benefits of lateral flow immunoassay (LFIA), such as operational simplicity, speed, and accuracy, we engineered AgPd nanocomposites with antibodies targeting GCA and AFP. Under the optimized conditions, the visual detection limit for GCA was established at 50 ng mL-1 and the cut-off value at 104 ng mL-1. And for AFP, the visual detection limit was 0.1 ng mL-1 and the cut-off value was 500 ng mL-1. The accuracy and feasibility of the strips were validated through the detection of 39 actual serum samples. The results highlight the potential of LFIA as a rapid and effective tool for clinical diagnosis. The developed LFIA method not only demonstrates accuracy and feasibility but also presents a promising avenue for the early diagnosis of hepatocellular carcinoma.


Subject(s)
Carcinoma, Hepatocellular , Liver Neoplasms , Humans , alpha-Fetoproteins , Carcinoma, Hepatocellular/diagnosis , Carcinoma, Hepatocellular/pathology , Liver Neoplasms/diagnosis , Liver Neoplasms/pathology , Biomarkers, Tumor , Glycocholic Acid , Immunoassay/methods
12.
J Mol Neurosci ; 72(3): 482-495, 2022 Mar.
Article in English | MEDLINE | ID: mdl-34623606

ABSTRACT

Mesenchymal stem cells (MSCs)-derived exosomes play significant roles in alleviating spinal cord injury (SCI). Previous study showed that long non-coding RNA tectonic family member 2 (TCTN2) was able to relieve SCI. Herein, whether TCTN2 exerted its roles in functional recovery after SCI via exosomes derived from MSCs was explored. The SCI model was established in rats, and the neurological function was evaluated using the Basso, Beattie, and Bresnahan (BBB) scoring. Lipopolysaccharide (LPS)-induced differentiated PC12 cells were used as an in vitro model for neurotoxicity research. The expression of genes and proteins was detected by qRT-PCR and Western blot. Exosomes were isolated by ultracentrifugation and qualified by TEM and Western blot. In vitro assays were performed using CCK-8 assay, EdU assay, and flow cytometry, respectively. Dual-luciferase reporter assay and RIP assay were used to confirm the target relationship between miR-329-3p and TCTN2 or insulin-like growth factor1 receptor (IGF1R). TCTN2 expression was down-regulated in SCI model rat and lipopolysaccharide (LPS)-stimulated PC12 cells. MSCs produced exosomes and could package TCTN2 into secreted exosomes. Tail vein injection of TCTN2 exosomes into rats significantly improved functional recovery of SCI. Meanwhile, TCTN2 exosomes treatment alleviated LPS-induced neuronal apoptosis, inflammation, and oxidative stress in vitro. Additionally, TCTN2 targeted miR-329-3p and subsequently regulated the expression of its target IGF1R. Rescue assays suggested that miR-329-3p/IGF1R axis mediated the beneficial effects of TCTN2 exosomes on LPS-treated PC12 cells. In all, exosomes derived from TCTN2-modified MSCs could improve functional recovery of SCI in vivo and attenuate LPS-induced neuronal apoptosis, inflammation, and oxidative stress in vitro via miR-329-3p/IGF1R axis, suggesting a novel insight into the development of MSC-exosomes-based therapy for SCI.


Subject(s)
Exosomes , Mesenchymal Stem Cells , MicroRNAs , RNA, Long Noncoding , Spinal Cord Injuries , Animals , Apoptosis/genetics , Exosomes/metabolism , Mesenchymal Stem Cells/metabolism , MicroRNAs/genetics , MicroRNAs/metabolism , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Rats , Receptor, IGF Type 1 , Spinal Cord/metabolism , Spinal Cord Injuries/genetics , Spinal Cord Injuries/metabolism , Spinal Cord Injuries/therapy
13.
J Hazard Mater ; 435: 129082, 2022 08 05.
Article in English | MEDLINE | ID: mdl-35650752

ABSTRACT

Multimodal lateral flow immunoassay (LFIA) has displayed its potential to improve practicability and elasticity of point-of-care testing. Herein, multifunctional core-shell-shell Au@Pt@Ag NPs loaded with dual-layer Raman reporter molecules of 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) with a characteristic combination of color-photothermal-Raman performance were constructed for colorimetric LFIA (CM-LFIA), photothermal LFIA (PT-LFIA) and surface-enhanced Raman scattering-based LFIA (SERS-LFIA), respectively. The highly specific nanoprobes, being obtained through the combination of the resulted dual-layer DTNB modified Au@Pt@Ag NPs with the antibody, were triumphantly utilized in exploring multimodal LFIA with one visual qualitative and two optional quantitative modes with excellent sensing sensitivity. Under optimal conditions, the limit of detection (LOD) for the model hazardous analyte dehydroepiandrosterone (DHEA) were 1.0 ng mL-1 for CM-LFIA, 0.42 ng mL-1 for PT-LFIA, and 0.013 ng mL-1 for SERS-LFIA, three of which were over 100-fold, 200-fold and 7 000-fold more sensitive than conventional visual AuNPs-based LFIA, respectively. In addition, the quantitative PT-LFIA and SERS-LFIA sensors worked well in spiked real samples with acceptable recoveries of 96.2 - 106.7% and 98.2 - 105.2%, respectively. This assay demonstrated that the developed multimodal LFIA had a great potential to be a powerful tool for accurate tracing hazardous analytes in complex samples.


Subject(s)
Gold , Metal Nanoparticles , Dithionitrobenzoic Acid , Immunoassay/methods , Spectrum Analysis, Raman/methods
14.
J Agric Food Chem ; 69(46): 13691-13699, 2021 Nov 24.
Article in English | MEDLINE | ID: mdl-34783242

ABSTRACT

Currently, the infection with Helicobacter pylori affects about half of the world's population, and the most common therapy to treat H. pylori is the first line clarithromycin-based triple therapy or the quadruple therapy. However, drug resistance, eradication in a low level, high rate of reinfection, and gastrointestinal side effects among the causative organisms for H. pylori infection pose a critical challenge to the global health care community. Therefore, new approaches to treat H. pylori infections are urgently needed. Chicken egg yolk constituting a source of immunoglobulin Y (IgY) has attracted noticeable attention for its advantages of cost-effective extraction, minimization of animal harm and suffering, and induction of no specific resistance and is, therefore, being regarded as an alternative therapy for H. pylori infection. This review is intended to summarize various H. pylori antigens for IgY preparation in terms of their application, mechanism, and limitations.


Subject(s)
Helicobacter pylori , Animals , Antibodies , Egg Yolk , Immunoglobulins , Urease
15.
Anal Methods ; 13(9): 1164-1171, 2021 03 11.
Article in English | MEDLINE | ID: mdl-33599661

ABSTRACT

Progesterone (P4) belongs to a factor that affects stress response and is a potential carcinogen, and saliva levels are expected to be a standard measurement for clinical diagnosis. In this study, a new type of nanoflower with both recognition functionality and catalytic substrate ability was prepared by copper phosphate, Pt/IrO2 nanocomposites (Pt/IrO2 NPs), streptavidin (SA) and horseradish peroxidase (HRP) via a one-pot co-precipitation strategy. Due to the enhanced catalytic activity and stability of Pt/IrO2@SA@HRP nanoflowers, we developed a powerful and sensitive multiple-catalysis ELISA to monitor progesterone in saliva. Multiple-catalysis ELISA based on a specific antibody and Pt/IrO2@SA@HRP nanoflowers exhibited a linear interval range from 0.217 ng mL-1 to 7.934 ng mL-1. The median inhibitory concentration (IC50) for progesterone is 1.311 ng mL-1 and the limit of detection (LOD = IC10) is 0.076 ng mL-1 in the proposed method. Satisfactory recoveries were in a range of 79.6-107% with an acceptable coefficient of variation (below 10.6%). Results of the multiple-catalysis ELISA and LC-MS/MS had a good coincidence. Our result unraveled that multiple-catalysis ELISA is a potentially serviceable tool for the detection of progesterone in saliva.


Subject(s)
Colorimetry , Progesterone , Chromatography, Liquid , Horseradish Peroxidase , Iridium , Nanostructures , Platinum , Saliva , Streptavidin , Tandem Mass Spectrometry
16.
Exp Ther Med ; 21(6): 646, 2021 Jun.
Article in English | MEDLINE | ID: mdl-33968177

ABSTRACT

Intervertebral disc degeneration and resulting low back pain arises from the programmed apoptosis of nucleus pulposus cells (NPCs). Recent studies show that hypoxia-inducible factor-1α plays a vital role in the etiology and pathogenesis of disc degeneration. However, the underlying mechanism of HIF-1α in NPCs is unclear. The present study identified 994 significant differentially expressed miRNAs by analyzing microarray data downloaded from the Gene Expression Omnibus database. MicroRNA(miR)-32-5p expression was 2.81-fold upregulated in NPCs compared with that of the healthy control tissues (P<0.05). A total of 331 significant differentially expressed mRNAs were identified, and PTEN was downregulated in NPCs of non-degenerative disc tissues from young patients. miR-32-5p was predicted to target the PTEN 3'-untranslated region (UTR). To confirm these results, in-vitro experiments investigating the molecular function of miR-32-5p and PTEN were performed. Furthermore, hypoxia induced miR-32-5p and PTEN expression. HIF-1α inhibited NPC proliferation and promoted cell apoptosis by regulating miR-32-5p and PTEN. miR-32-5p promoted NPC proliferation and decreased cell apoptosis. Next, it was verified whether miR-32-5p targeted the PTEN 3'-UTR using dual-luciferase reporter assays. Finally, it was observed that PI3K/AKT/mTOR signaling pathway was upregulated by a miR-32-5p mimic, which improved cell proliferation and decreased apoptosis. Importantly, PTEN was downregulated in these experiments; and inhibition of miR-32-5p had the opposite effect. Overall, these results demonstrate that HIF-1α regulates cell proliferation and apoptosis by controlling the miR-32-5p/PTEN/PI3K/AKT/mTOR axis in NPCs.

17.
J Chromatogr A ; 1503: 1-11, 2017 Jun 23.
Article in English | MEDLINE | ID: mdl-28499596

ABSTRACT

Microcystins (MCs) are cyclic heptapeptide toxins and tumor promoters produced by cyanobacteria, which threaten the health of humans. In this study, magnetic porous ß-cyclodextrin polymer (Fe3O4@SiO2@P-CDP) was synthesized and characterized by transmission electron microscopy, scanning electron microscopy, energy dispersive X-ray spectrometry, Fourier transform infrared spectrometry, X-ray diffraction, nitrogen adsorption porosimetry and vibrating sample magnetometer. The synthesized Fe3O4@SiO2@P-CDP particles were then used for magnetic solid-phase extraction (MSPE) of MCs from environmental water samples, and exhibited excellent extraction performance, especially for MC-RR. Coupled with high-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS), a simple, efficient and sensitive method for determination of trace levels of MCs was established. After the optimization of conditions, wide linear ranges (2.0-1000pgmL-1), good linearity (r2≥0.9996) and acceptable repeatability (RSD≤9.4%, n=5) were obtained. The limits of detection (LODs, S/N=3) and limits of quantification (LOQs, S/N=10) for three MCs (MC-LR, MC-RR and MC-YR) were in the range of 1.0-2.0pgmL-1 and 2.0-5.0pgmL-1, respectively. Typical water samples were analyzed by the developed method, and trace levels of MC-LR and MC-RR were detected. The results demonstrate that the developed method has great potential for the determination of MCs in complicated matrix.


Subject(s)
Environmental Monitoring/methods , Magnetics , Microcystins/isolation & purification , Solid Phase Extraction/methods , beta-Cyclodextrins/chemistry , Adsorption , Cellulose/chemistry , Cyanobacteria/chemistry , Cyclodextrins/chemistry , Limit of Detection , Microcystins/analysis , Silicon Dioxide/chemistry , X-Ray Diffraction
18.
J Chromatogr A ; 1443: 54-61, 2016 Apr 22.
Article in English | MEDLINE | ID: mdl-27033983

ABSTRACT

A simple, rapid and sensitive method for determination of trace levels of domoic acid (DA) in seawater was developed, based on a magnetic solid-phase extraction (MSPE) followed by high-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS). Five kinds of ferrite magnetic nanospheres (MFe2O4; M=Fe, Co, Ni, Cu and Zn) were prepared and first used as sorbents for MSPE of DA and removal of salt interference. Under the same extraction and elution conditions, CuFe2O4 magnetic nanospheres provided the best pretreatment performance, which were then characterized in detail. After further optimization of conditions, the developed method showed good linearity (r(2)=0.9991) with the range of 5-1000 pg mL(-1), low limit of detection (2.5 pg mL(-1); S/N=3:1), low limit of quantification (5.0 pg mL(-1); S/N=10:1), and good recoveries (86.0-98.1%) with acceptable repeatability (RSD ≤ 6.5%; n=3) in seawater samples. The results demonstrated that the ferrite magnetic nanospheres are promising sorbents for efficient extraction of highly polar analytes from high ionic strength solutions.


Subject(s)
Chromatography, Liquid , Environmental Monitoring/methods , Kainic Acid/analogs & derivatives , Magnetics , Nanospheres/chemistry , Solid Phase Extraction/methods , Tandem Mass Spectrometry , Ferric Compounds/chemistry , Kainic Acid/analysis , Seawater/chemistry
19.
Biosens Bioelectron ; 68: 475-480, 2015 Jun 15.
Article in English | MEDLINE | ID: mdl-25621999

ABSTRACT

Recently we demonstrated oriented formation of gold nanoparticle (AuNP) dimers for ultrasensitive sensing oligonucleotides (J. Am. Chem. Soc. 2013, 135, 12338). Herein, we investigate the reverse process of this sensing mechanism using target analytes to disassemble the orient-aggregated AuNP dimers. This enables us to expand the analytes from oligonucleotides to other molecules, e.g. highly sensitive and selective determination of microcystin-LR (MC-LR) is selected for a demonstration in this work. Aptamers specific to the target molecules are used as linkers to prepare the AuNP dimers. In the presence of the target molecule, the aptamer changes its structure to bind the target molecule. Thus the pre-formed AuNP dimers are disassembled. As a result, the solution color is changed from blue to red. This sensing design retains the advantages of the previously developed sensors based on target molecules guided formation of AuNP dimers, e.g. the overwhelming sensitivity and stability comparing with those non-oriented sensors based on the formation of large aggregates, with the additional advantages as follows: 1) the target molecules are expanded from oligonucleotides to arbitrary molecules that can specifically bind to aptamers; 2) the color change is completed within 5 min, while the previous sensor based on the formation of AuNP dimers cost ~1 hour to obtain stable responses.


Subject(s)
Biosensing Techniques , Microcystins/isolation & purification , Aptamers, Nucleotide/chemistry , Colorimetry , Gold/chemistry , Humans , Marine Toxins , Metal Nanoparticles/chemistry , Microcystins/chemistry
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