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In this study, a novel colorimetric and fluorescent dual-mode ELISA based on glucose oxidase (GOx)-triggered Fenton reaction was developed for the qualitative and quantitative detection of danofloxacin (DAN). In this system, streptavidin-linked biotinylated anti-DAN-monoclonal antibody (SA-Bio-mAb) and biotinylated GOx (Bio-GOx) form the immune complex mAb-Bio-SA-Bio-GOx. In the absence of DAN, the mAb-Bio-SA-Bio-GOx would be immobilized by combining with coated DAN-BSA and catalyzed glucose to generate H2O2. The Fenton reaction between H2O2 and Fe2+ generated hydroxyl radicals, which oxidized the o-phenylenediamine to 2,3-diamino-phenazine. A dual-signal immunoassay with colorimetry and fluorescence as the signal readout was established. In the presence of DAN, DAN and DAN-BSA competed with Bio-mAb, decreasing the connection between immune complexes and DAN-BSA and finally resulting in lower signal of colorimetry and fluorescence. Under optimal conditions, the limit of detection of the fluorescence immunoassay was 0.337 ng/mL and was 5.24-fold lower than that of traditional ELISA. The colorimetric immunoassay cut-off value was 30 ng/mL in milk. The average recoveries of the method for milk samples that are spiked with different concentrations of DAN were 91.1 to 128.3%, with a coefficient of variation of 0.7 to 8.2%. These results of the method exhibited good agreement with those of liquid chromatography-tandem mass spectrometry system (LC-MS/MS) method. In brief, this work provides an improved screening strategy with high sensitivity and accuracy for the qualitative or quantitative detection of DAN in milk monitoring.
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Fluoroquinolonas/análise , Glucose Oxidase/química , Imunoensaio/veterinária , Leite/química , Animais , Bovinos , Colorimetria/métodos , Colorimetria/veterinária , Fluorescência , Peróxido de Hidrogênio/química , Imunoensaio/métodos , Testes Imunológicos/veterinária , Limite de DetecçãoRESUMO
Pathogens, mycotoxins, or antibiotics may exist in a food sample. Micro- and macromolecular substances must be detected quickly. A rapid and convenient lateral flow immunoassay (LFI) integrated with competitive and sandwich models was developed to detect micro- and macromolecular substances. In this study, aflatoxin M1 (AFM1) and Escherichia coli O157:H7 were selected as the micro- and macromolecular substances, respectively. Two test lines in the LFI test strip were evaluated to detect AFM1 and E. coli O157:H7 by competitive and sandwich models. Results showed that the limits of detection for detecting AFM1 and E. coli O157:H7 were 50 pg·mL-1 and 1.58 × 104 cfu·mL-1, respectively. The whole assay time was 30 min. The recoveries of gold nanoparticle-LFI ranged from 78.0 to 111.6% with coefficients of variation in the range of 3.9 to 8.5% for the detection of AFM1. For the detection of E. coli O157:H7, the range of recoveries was from 70.1 to 89.6% with coefficients of variation ranging from 4.9 to 13.0%. This study not only tested sensitivity and specificity, but also was a systematic study of location of 2 test lines of the LFI test strip integrated with competitive and sandwich models.
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Aflatoxina M1/isolamento & purificação , Escherichia coli O157/isolamento & purificação , Imunoensaio/métodos , Leite/química , Leite/microbiologia , Animais , Microbiologia de Alimentos , Ouro , Nanopartículas MetálicasRESUMO
PURPOSE: This study aimed to develop and validate accessible artificial neural network and decision tree models to predict the risk of lower limb lymphedema after cervical cancer surgery. METHODS: We selected 759 patients who underwent cervical cancer surgery at the Hunan Cancer Hospital from January 2010 to January 2020, collecting demographic, behavioral, clinicopathological, and disease-related data. The artificial neural network and decision tree techniques were used to construct prediction models for lower limb lymphedema after cervical cancer surgery. Then, the models' predictive efficacies were evaluated to select the optimal model using several methods, such as the area under the receiver operating characteristic curve and accuracy, sensitivity, and specificity tests. RESULTS: In the training set, the artificial neural network and decision tree model accuracies for predicting lower limb lymphedema after cervical cancer surgery were 99.80% and 88.14%, and the sensitivities 99.50% and 74.01%, respectively; the specificities were 100% and 95.20%, respectively. The area under the receiver operating characteristic curve was 1.00 for the artificial neural network and 0.92 for the decision tree model. In the test set, the artificial neural network and decision tree models' accuracies were 86.70% and 82.02%, and the sensitivities 65.70% and 67.11%, respectively; the specificities were 96.00% and 89.47%, respectively. CONCLUSION: Both models had good predictive efficacy for lower limb lymphedema after cervical cancer surgery. However, the predictive performance and stability were superior in the artificial neural network model than in the decision tree model.
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Background: To analyze the factors influencing lower limb lymphedema (LLL) after cervical cancer surgery and provide a scientific reference for its effective prevention and control. Methods and Results: Patients at a tertiary tumor hospital between January 2010 and January 2020 who received surgical treatment for cervical cancer were included in this study. Patients were divided into lymphedema case group (n = 253) and control group (n = 506) according to lymphedema occurrence, and univariate and multivariate analyses were used to analyze the factors influencing the occurrence of LLL after cervical cancer surgery. Multifactor conditional logistic regression analysis revealed that risk factors for lymphedema occurrence included body mass index, level of education, the presence of preoperative radiotherapy and chemotherapy, radiotherapy within 3 months after postoperative chemoradiotherapy, emergence of coronary heart disease within 3 months, vaginal disease, occurrence of postoperative complications, cervical cancer diagnosis before the manifestation of menstrual abnormalities, and a history of previous surgery. Conclusion: Postoperative LLL after cervical cancer surgery is a chronic progressive disease, and no cure for LLL has been identified. Thus, determining the risk factors associated with LLL occurrence after uterine and cervical cancer surgery and the development of targeted prevention and early intervention strategies is urgently needed.
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Linfedema , Neoplasias do Colo do Útero , Feminino , Humanos , Estudos de Casos e Controles , Neoplasias do Colo do Útero/patologia , Extremidade Inferior/patologia , Estudos Retrospectivos , Excisão de Linfonodo/efeitos adversos , Linfedema/etiologia , Fatores de RiscoRESUMO
Objectives: The aim of this study was to find a new loss function to automatically segment temporal lobes on localized CT images for radiotherapy with more accuracy and a solution to dealing with the classification of class-imbalanced samples in temporal lobe segmentation. Methods: Localized CT images for radiotherapy of 70 patients with nasopharyngeal carcinoma were selected. Radiation oncologists sketched mask maps. The dataset was randomly divided into the training set (n = 49), the validation set (n = 7), and the test set (n = 14). The training set was expanded by rotation, flipping, zooming, and shearing, and the models were evaluated using Dice similarity coefficient (DSC), Jaccard similarity coefficient (JSC), positive predictive value (PPV), sensitivity (SE), and Hausdorff distance (HD). This study presented an improved loss function, focal generalized Dice-binary cross-entropy loss (FGD-BCEL), and compared it with four other loss functions, Dice loss (DL), generalized Dice loss (GDL), Tversky loss (TL), and focal Tversky loss (FTL), using the U-Net model framework. Results: With the U-Net model based on FGD-BCEL, the DSC, JSC, PPV, SE, and HD were 0.87 ± 0.11, 0.78 ± 0.11, 0.90 ± 0.10, 0.87 ± 0.13, and 4.11 ± 0.75, respectively. Except for the SE, all the other evaluation metric values of the temporal lobes segmented by the FGD-BCEL-based U-Net model were improved compared to the DL, GDL, TL, and FTL loss function-based U-Net models. Moreover, the FGD-BCEL-based U-Net model was morphologically more similar to the mask maps. The over- and under-segmentation was lessened, and it effectively segmented the tiny structures in the upper and lower poles of the temporal lobe with a limited number of samples. Conclusions: For the segmentation of the temporal lobe on localized CT images for radiotherapy, the U-Net model based on the FGD-BCEL can meet the basic clinical requirements and effectively reduce the over- and under-segmentation compared with the U-Net models based on the other four loss functions. However, there still exists some over- and under-segmentation in the results, and further improvement is needed.
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A signal-enhanced LFIA based on tyramine (TYR)-induced AuNPs aggregation has been developed for the sensitive detection of danofloxacin (DAN). In the model, the hydroxyl radical produced by HRP catalyzing H2O2 can trigger the TYR-AuNPs to aggregate on the T or C line for enhancing the detection signal. The linear range of TYR-AuNPs LFIA was 0.25-5 ng mL-1 with the limit of detection (LOD) of 0.032 ng mL-1, and the LOD was 8-fold lower than that of the traditional AuNPs LFIA (0.26 ng mL-1). The TYR-AuNPs LFIA could be used with the naked eyes to qualitatively detect DAN with a cut-off limit of 2.5 ng mL-1, which was 4-fold lower than that of the traditional AuNPs LFIA (10 ng mL-1). The recoveries of TYR-AuNPs LFIA were 86.04-105.14% and 92.41-110.19%, with the coefficient of variation of 1.71-2.05% and 4.42-5.89% in chicken and pork, respectively.
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Ouro , Nanopartículas Metálicas , Fluoroquinolonas , Peróxido de Hidrogênio , Imunoensaio , Limite de Detecção , TiraminaRESUMO
Objective: To explore the performance of Multi-scale Fusion Attention U-Net (MSFA-U-Net) in thyroid gland segmentation on localized computed tomography (CT) images for radiotherapy. Methods: We selected localized radiotherapeutic CT images from 80 patients with breast cancer or head and neck tumors; label images were manually delineated by experienced radiologists. The data set was randomly divided into the training set (n = 60), the validation set (n = 10), and the test set (n = 10). We expanded the data in the training set and evaluated the performance of the MSFA-U-Net model using the evaluation indices Dice similarity coefficient (DSC), Jaccard similarity coefficient (JSC), positive predictive value (PPV), sensitivity (SE), and Hausdorff distance (HD). Results: For the MSFA-U-Net model, the DSC, JSC, PPV, SE, and HD values of the segmented thyroid gland in the test set were 0.90 ± 0.09, 0.82± 0.11, 0.91 ± 0.09, 0.90 ± 0.11, and 2.39 ± 0.54, respectively. Compared with U-Net, HRNet, and Attention U-Net, MSFA-U-Net increased DSC by 0.04, 0.06, and 0.04, respectively; increased JSC by 0.05, 0.08, and 0.04, respectively; increased SE by 0.04, 0.11, and 0.09, respectively; and reduced HD by 0.21, 0.20, and 0.06, respectively. The test set image results showed that the thyroid edges segmented by the MSFA-U-Net model were closer to the standard thyroid edges delineated by the experts than were those segmented by the other three models. Moreover, the edges were smoother, over-anti-noise interference was stronger, and oversegmentation and undersegmentation were reduced. Conclusion: The MSFA-U-Net model could meet basic clinical requirements and improve the efficiency of physicians' clinical work.
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Danofloxacin (DAF), a third-generation fluroquinolone (FQ), is widely used as a broad-spectrum antibacterial drug to prevent diseases in livestock and poultry. In this study, a highly specific and sensitive monoclonal antibody (mAb) against DAF was prepared. Also, the mAb was used for the indirect competitive enzyme-linked immunosorbent assay (icELISA) and immunochromatographic strip for the detection of DAF residues in meat. The IC50 of the icELISA based on this mAb was 1.39 ng/mL, and the limit of detection was 0.2 ng/mL. According to the cross-reactivity (CR) experiment, the ELISA that we developed was highly specific and had low CR with other FQ analogues. Moreover, the cut-off of the immunochromatographic strip developed for detecting DAF in meat was 5 ng/mL. Overall, the developed ELISA and immunochromatographic strip based on the prepared mAb were proved reliable for the rapid detection of DAF in meat and can be considered as effective screening methods for food safety and quality management.
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Fluoroquinolones are synthetic antibiotics that are commonly used in animal husbandry, and the consumption of animal products with fluoroquinolone residues has imposed a serious threat to human health. Here, we report a plasmonic enzyme-linked immunosorbent assay (pELISA) method based on oxidative etching of silver nanoprisms (AgNPRs) for the quantitative and qualitative detection of danofloxacin (DAN), a fluoroquinolone antibiotic. AgNPRs that undergo colorimetric changes upon oxidative etching by H2O2 serve as the signal transducer in our design. An indirect competitive pELISA was constructed by introducing biotinylated monoclonal antibody (mAb), streptavidin and biotinylated glucose oxidase, which catalyzes the generation of H2O2 for etching AgNPRs. The quantitative detection limit of the proposed method was 0.24 ng mL-1 for DAN. The qualitative detection limit for DAN reached 0.32 ng mL-1, which was 32-fold lower than that of the assay using 3,3',5,5'-tetramethylbenzidine (TMB) as the signal transducer. The average recoveries of DAN in milk ranged from 103% to 121%, with a coefficient of variation of 0.6-3.41%. The recovery results were further confirmed using liquid chromatography-tandem mass spectrometry. In summary, the proposed AgNPR-etching pELISA exhibits high sensitivity, good accuracy and excellent reliability for the quantitative and qualitative detection of DAN in milk.
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Ensaio de Imunoadsorção Enzimática , Fluoroquinolonas/análise , Nanopartículas Metálicas/química , Prata/química , Animais , Biotina/química , Biotina/metabolismo , Glucose Oxidase/química , Glucose Oxidase/metabolismo , Leite/química , Tamanho da Partícula , Propriedades de SuperfícieRESUMO
In this study, we developed a competitive colorimetric immunoassay for qualitative detection of DAN based on oxidation of iron (â ¡) (Fe2+) in the presence of glucose oxidase (GOx) and color change induced by Fe2+-phenanthroline (Phen) chromogenic system. Streptavidin (SA) acted as a linker between biotinylated anti-DAN-monoantibody (bio-mAb) and biotinylated GOx (bio-GOx) to form the immunocomplexes bio-mAb-SA-bio-GOx. In the absence of DAN, the immunocomplexes bio-mAb-SA-bio-GOx combining with coated DAN-ovalbumin (DAN-OVA) will be immobilized and catalyze glucose to produce H2O2. Fe2+ is oxidized to Fe3+ by H2O2, giving rise to a colorless result. In the presence of DAN, Fe2+ produces a chelation reaction with Phen, leading to orange-red color. Under optimal conditions, the detection limit (LOD) by naked eyes was 2.5 ng mL-1 in milk, chicken, beef, and pork samples. Low LOD, no matrix effect, and no signal reader requirement make it possibly applied to quickly screen DAN on site.
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Técnicas Biossensoriais/métodos , Colorimetria/métodos , Fluoroquinolonas/análise , Glucose Oxidase/metabolismo , Imunoensaio/métodos , Quelantes de Ferro/química , Fenantrolinas/química , Biocatálise , Catálise , Enzimas Imobilizadas/química , Enzimas Imobilizadas/metabolismo , Fluoroquinolonas/química , Glucose/química , Glucose Oxidase/química , Peróxido de Hidrogênio/química , Limite de DetecçãoRESUMO
Escherichia coli O157:H7 (E. coli O157:H7) is a potential threat to human health; thus, a rapid and sensitive method for detecting it is necessary. We designed a single-stranded DNA that contained an appended block and anchoring block. The appended block acted as a scaffold to prepare fluorescent Ag nanoclusters (AgNCs). The anchoring block contained Poly A, which bound with the surface of gold nanoparticles to quench the fluorescence of AgNCs. An interesting ELISA approach for detecting E. coli O157:H7 was established via fluorescent quenching of DNA-stabilized AgNCs by using a sandwich complex. The changes in fluorescence intensity of AgNCs were used to quantitatively detect E. coli O157:H7. The sensitivity for detecting E. coli O157:H7 reached 1.905â¯×â¯103â¯CFU/mL with a good linear range. Compared with conventional ELISA, the sensitivity of this technique increased by 30-fold. Moreover, this method demonstrated specificity and reproducibility and could be used in food samples.
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Ensaio de Imunoadsorção Enzimática , Escherichia coli O157/isolamento & purificação , Nanopartículas Metálicas/química , Prata/química , Animais , DNA Bacteriano/isolamento & purificação , Fluorescência , Contaminação de Alimentos/análise , Microbiologia de Alimentos , Ouro/química , Leite/microbiologia , Sensibilidade e EspecificidadeRESUMO
Fluorescent microsphere (FM) is widely used as probe in immunochromatographic assay (ICA). However, the performance of conventional FM is limited because of the aggregation-caused quenching effect. Herein, we compared a kind of conventional FM (DMFFM, loading DMF) with novel aggregation-induced emission FM (AIEFM, loading TCBPE). The fluorescence intensity of DMFFM initially increased and then decreased as the concentrations of the loading DMF increased. The fluorescence intensity of AIEFM increased as the concentrations of the loading TCBPE increased and retained a high value. AIEFM was compared with two commercial FMs purchased from Ocean (OFM) and Merk (MFM). The maximum fluorescence intensity and relative quantum yield of AIEFM was approximately 5 and 4.5 times higher than those of two commercial FMs. We used the novel AIEFM as a probe to improve the sensitivity of ICA. When Escherichia coli O157:H7 was detected as the target, the limit of detection of ICA based on AIEFM, OFM and MFM were 3.98â¯×â¯103â¯CFU/mL, 4.48â¯×â¯104 and 2.78â¯×â¯104â¯CFU/mL, respectively. The ICA of AIEFM had 11 and 7 times improvement in sensitivity compared with that of OFM and MFM. Our results could be used as a basis for novel probes in practical ICA applications.
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Escherichia coli O157/isolamento & purificação , Corantes Fluorescentes/química , Imunoensaio/instrumentação , Técnicas Biossensoriais/instrumentação , Dimerização , Infecções por Escherichia coli/microbiologia , Fluorescência , Humanos , MicroesferasRESUMO
There are conflicting records on the identity of Chinese medical herbs, and it is known that different plant materials are used under the same common names in different regions in China. However, there is no study on the genetic heterogeneity of medical herbs in any market outside of China. In this report, Chinese medical herbs under common names Radix Quinquefolii (American Ginseng or Xiyangshen), Radix Astragali (Huangqi), Radix Notoginseng (Tianqi), Coxtex Cinnamomum (Guipi), Radix Isatidis (Banlangen), Radix Codonopsis (Dangshen) and Radix Rehmannia (Shengdi) were collected from three independent herbal shops in Singapore and their DNAs were isolated and subjected to fluorescence Amplified Fragment Length Polymorphism (AFLP) analysis. While samples for Radix Quinquefolii and Radix Astragali were homogenous genetically [similarity index (SI) = 0.85-1.00] across the three shops, genetic heterogeneity was found for the other herbs (SI < 0.7). For example, four samples of Radix Codonopsis were of three distinct patterns (SI < 0.6). Our results highlight the situation that genetically distinct herbal materials are labeled and marketed under the same common names in an international market of Chinese medical herbs, which may contribute to inconsistency in quality and efficacy.
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Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/normas , Citometria de Fluxo , Panax/química , Polimorfismo de Fragmento de Restrição , China , Estudos de Avaliação como Assunto , Citometria de Fluxo/métodos , Humanos , Extratos Vegetais/química , Extratos Vegetais/normas , Controle de Qualidade , Especificidade da EspécieRESUMO
OBJECTIVE: We aimed to investigate the effects of progesterone on gene expression and function of both myometrium and circulating leukocytes. METHODS: We recruited women participating in a randomized clinical trial of progesterone to prevent preterm delivery. These participants had a twin pregnancy and were managed in 1 of 2 tertiary referral centers. Participants were treated with progesterone (90 mg vaginally) or placebo from 24 to 34 weeks of pregnancy. The outcome measures were myometrial and leukocyte gene expression and expression of cell surface markers in circulating leukocytes, all quantified ex vivo. RESULTS: Prolonged in vivo administration of progesterone inhibited myometrial expression of connexins 26 and 43, endothelial nitric acid synthase (eNOS), and the prostaglandin receptor EP2 ex vivo. Administration of progesterone also increased numbers of circulating neutrophils while decreasing lymphocyte proportions and decreasing neutrophil CD11b expression. CONCLUSION: The observed effects of prolonged in vivo administration of progesterone will minimize the ability of the uterus to contract as a synctium and the ability of peripheral blood leukocytes to migrate into the myometrium during parturition. We suggest that these are putative mechanisms by which progesterone might prevent preterm birth in women at high risk.
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Leucócitos Mononucleares/metabolismo , Miométrio/metabolismo , Gravidez Múltipla/metabolismo , Nascimento Prematuro/metabolismo , Progesterona/administração & dosagem , Progesterona/sangue , Adulto , Biomarcadores/sangue , Antígeno CD11b/metabolismo , Células Cultivadas , Estudos de Coortes , Conexina 43/antagonistas & inibidores , Conexina 43/metabolismo , Estradiol/sangue , Feminino , Regulação da Expressão Gênica , Humanos , Hidrocortisona/sangue , Leucócitos Mononucleares/efeitos dos fármacos , Miométrio/efeitos dos fármacos , Gravidez , Complicações na Gravidez/tratamento farmacológico , Complicações na Gravidez/metabolismo , Complicações na Gravidez/prevenção & controle , Gravidez Múltipla/efeitos dos fármacos , Nascimento Prematuro/tratamento farmacológico , Nascimento Prematuro/prevenção & controle , Fatores de TempoRESUMO
MICs of piperacillin/tazobactam are conventionally determined by varying the concentration of piperacillin in the presence of a fixed 4 mg/L tazobactam. When tested in this way, the MIC distribution for Klebsiella isolates with extended-spectrum beta-lactamases (ESBLs) is strongly bimodal, such that many producers are inhibited at 16 + 4 mg/L whilst others require MICs of > or =512 + 4 mg/L. When, however, piperacillin/tazobactam was tested as a fixed 8:1 ratio, the MIC distribution became unimodal. If clavulanate 4 mg/L was combined with piperacillin, a unimodal MIC distribution was seen for ESBL-producing Klebsiella spp. but a bimodal distribution arose if the clavulanate concentration was reduced to 0.25 mg/L. These data for alternative combinations suggested that the bimodal MIC distribution seen for piperacillin + tazobactam 4 mg/L was a titration effect, not a reflection of some ESBLs being resistant to tazobactam. Even within single strains, as defined by serotype and DNA fingerprints, there was considerable variation in susceptibility to piperacillin + tazobactam 4 mg/L, with some representatives highly susceptible and others highly resistant. Some of the more resistant representatives produced more of their ESBL, or had a greater number of beta-lactamase types, but these associations were not universal. Elevated resistance to piperacillin + tazobactam was not associated with porin change in any ESBL producer examined, but has been found by others.