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Protein phosphorylation, one of the main protein post-translational modifications, is required for regulating various life activities. Kinases and phosphatases that regulate protein phosphorylation in humans have been targeted to treat various diseases, particularly cancer. High-throughput experimental methods to discover protein phosphosites are laborious and time-consuming. The burgeoning databases and predictors provide essential infrastructure to the research community. To date, >60 publicly available phosphorylation databases and predictors each have been developed. In this review, we have comprehensively summarized the status and applicability of major online phosphorylation databases and predictors, thereby helping researchers rapidly select tools that are most suitable for their projects. Moreover, the organizational strategies and limitations of these databases and predictors have been highlighted, which may facilitate the development of better protein phosphorylation predictors in silico.
Assuntos
Proteínas Quinases , Processamento de Proteína Pós-Traducional , Humanos , Fosforilação , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteínas/metabolismo , Bases de Dados de ProteínasRESUMO
Objective: To evaluate the clinical efficacy of vestibular assessment and rehabilitation training in patients with peripheral vestibular vertigo. Method: This was a retrospective study. A total of 169 patients diagnosed with peripheral vestibular vertigo, admitted to Cangzhou People's Hospital between January 2020 and January 2023 were divided into control group (83 cases) and observation group (86 cases). The control group received medication-based treatment, while the observation group was provided with combined treatment of medication and vestibular rehabilitation training. Assessment of recovery included the Dizziness Handicap Inventory (DHI), Vestibular Symptom Index (VSI), and Activities-specific Balance Confidence (ABC) scale before and at two, four, and eight weeks post-treatment. Psychological status, sleep quality, and life quality were evaluated. Both groups underwent the Fukuda stepping test and timed balance test. Result: At two, four, and eight weeks post-treatment, both groups exhibited significantly lower DHI-P, DHI-F, DHI-E, VSI, and ABC scores compared to pre-treatment (p<0.05). The observation group showed significantly lower DHI-P, DHI-F, DHI-E, VSI, and ABC scores than the control group at two and four weeks post-treatment (p<0.05). After treatment, both groups demonstrated reduced body deviation angles and increased time without falling in the Fukuda stepping test (p<0.05). Notably, the observation group had significantly better outcomes (p<0.05). Conclusion: In comparison to medication-based treatment alone, a combined approach involving medication treatment and vestibular rehabilitation training may demonstrate early improvements in vertigo symptoms, enhance balance capabilities, and ameliorate psychological well-being, sleep quality, and overall quality of life for patients.
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The Adaptive Boosting (AdaBoost) algorithm is a widely used ensemble learning framework, and it can get good classification results on general datasets. However, it is challenging to apply the AdaBoost algorithm directly to imbalanced data since it is designed mainly for processing misclassified samples rather than samples of minority classes. To better process imbalanced data, this paper introduces the indicator Area Under Curve (AUC) which can reflect the comprehensive performance of the model, and proposes an improved AdaBoost algorithm based on AUC (AdaBoost-A) which improves the error calculation performance of the AdaBoost algorithm by comprehensively considering the effects of misclassification probability and AUC. To prevent redundant or useless weak classifiers the traditional AdaBoost algorithm generated from consuming too much system resources, this paper proposes an ensemble algorithm, PSOPD-AdaBoost-A, which can re-initialize parameters to avoid falling into local optimum, and optimize the coefficients of AdaBoost weak classifiers. Experiment results show that the proposed algorithm is effective for processing imbalanced data, especially the data with relatively high imbalances.
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Immunomagnetic separation (IMS) is an effective tool for the preconcentration and purification of food-borne pathogens from complex food samples because of its high capture efficiency (CE). In conventional IMS, antibodies are usually conjugated on the surface of magnetic beads (MB); the random orientation and conformation changes of antibodies on the MB surface can decrease their bioactivity. Moreover, the Brownian motion of immobilized antibodies is weakened, thereby rendering their binding efficiency with bacteria lower than that of free antibodies. Thus, abundant antibodies are commonly required to ensure high CE for IMS, particularly for large volumes. In this study, a 2-step large-volume magnetic separation (10 mL) was proposed to preconcentrate Listeria monocytogenes from pasteurized milk. First, the biotinylated anti-L. monocytogenes monoclonal antibodies (mAb) were bound with L. monocytogenes in 10 mL of diluted milk through an antigen-antibody interaction, and then streptavidin-labeled MB were used to capture biotin-mAb coated with L. monocytogenes by biotin and streptavidin interaction. Under optimal conditions, the CE of 2-step magnetic separation was >90% with L. monocytogenes concentrations ranging from 8 × 100 to 8 × 104 cfu/mL, whereas the amount of biotin-mAb was 14 fold lower than that of the conventional IMS method. Coupled with a PCR assay, the detection limit of the proposed method was 8 × 100 cfu/mL in pure culture and 8 × 101 cfu/mL in pasteurized milk without any pre-enrichment process. Moreover, the overall detection time, including sample preparation, large-volume magnetic separation, and PCR, took less than 7 h. In summary, the developed 2-step large-volume IMS combined with PCR was highly sensitive and low cost and, thus, has considerable potential for the rapid screening of food-borne pathogenic bacteria.
Assuntos
Microbiologia de Alimentos , Separação Imunomagnética/veterinária , Listeria monocytogenes/isolamento & purificação , Leite/microbiologia , Pasteurização , Reação em Cadeia da Polimerase/veterinária , Animais , Anticorpos Antibacterianos/isolamento & purificação , Biotina , Separação Imunomagnética/métodos , Listeria monocytogenes/imunologia , Reação em Cadeia da Polimerase/métodos , Sensibilidade e EspecificidadeRESUMO
Glutathione (GSH) plays important roles in the human body including protecting cells from oxidative damages and maintaining cellular redox homeostasis. Thus, developing a fast and sensitive method for detecting GSH levels in living bodies is of great importance. Many methods have been developed and used for GSH detection, such as high-performance liquid chromatography, capillary electrophoresis, and fluorescence resonance energy-based methods. However, these methods often lack sensitivity as well as efficiency. Herein, a rapid and sensitive method for glutathione detection was developed based on a fluorescence-enhanced "turn-on" strategy. In this study, a unique and versatile bifunctional linker 3-[(2-aminoethyl) dithio]propionic acid (AEDP)-modified gold nanoparticle (Au@PLL-AEDP-FITC) probe was designed for the simple, highly sensitive intracellular GSH detection, combined with the FRET technique. In the presence of GSH, the disulfide bonds of AEDP on Au@PLL-AEDP-FITC were broken through competition with GSH, and FITC was separated from gold nanoparticles, making the fluorescence signal switch to the "turn on" state. A change in the fluorescence signal intensity has a great linear positive correlation with GSH concentration, in the linear range from 10 nM to 180 nM (R2 = 0.9948), and the limit of detection (LOD) of 3.07 nM, which was lower than other reported optical nanosensor-based methods. Au@PLL-AEDP-FITC also has great selectivity for GSH, making it promising for application in complex biological systems. The Au@PLL-AEDP-FITC probe was also successfully applied in intracellular GSH imaging in HeLa cells with confocal microscopy. In short, the Au@PLL-AEDP-FITC probe-based fluorescence-enhanced "turn-on" strategy is a sensitive, fast, and effective method for GSH detection as compared with other methods. It can be applied in complex biological systems such as cell systems, with promising biological-medical applications in the future.
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Corantes Fluorescentes/química , Glutationa/análise , Transferência Ressonante de Energia de Fluorescência , Corantes Fluorescentes/síntese química , Ouro/química , Células HeLa , Humanos , Nanopartículas Metálicas/química , Propionatos/químicaRESUMO
Circulating tumor cell (CTC) detection and enumeration have been considered as a noninvasive biopsy method for the diagnosis, characterization, and monitoring of various types of cancers. However, CTCs are exceptionally rare, which makes CTC detection technologically challenging. In the past few decades, much effort has been focused on highly efficient CTC capture, while the activity of CTCs has often been ignored. Here, we develop an effective method for nondestructive CTC capture, release, and detection. Folic acid (FA), as a targeting molecule, is conjugated on magnetic nanospheres through a cleavable disulfide bond-containing linker (cystamine) and a polyethylene glycol (PEG2k) linker, forming MN@Cys@PEG2k-FA nanoprobes, which can bind with folate receptor (FR) positive CTCs specifically and efficiently, leading to the capture of CTCs with an external magnetic field. When approximately 150 and 10 model CTCs were spiked in 1 mL of lysis blood, 93.1 ± 2.9% and 80.0 ± 9.7% CTCs were recovered, respectively. In total, 81.3 ± 2.6% captured CTCs can be released from MN@Cys@PEG2k-FA magnetic nanospheres by treatment with dithiothreitol. The released CTCs are easily identified from blood cells for specific detection and enumeration combined with immunofluorescence staining with a limit of detection of 10 CTC mL-1 lysed blood. Moreover, the released cells remain healthy with high viability (98.6 ± 0.78%) and can be cultured in vitro without detectable changes in morphology or behavior compared with healthy untreated cells. The high viability of the released CTCs may provide the possibility for downstream proteomics research of CTCs; therefore, cultured CTCs were collected for proteomics. As a result, 3504 proteins were identified. In conclusion, the MN@Cys@PEG2k-FA magnetic nanospheres prepared in this study may be a promising tool for early-stage cancer diagnosis and provide the possibility for downstream analysis of CTCs.
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Cistamina/química , Ácido Fólico/química , Nanosferas/química , Células Neoplásicas Circulantes/patologia , Separação Celular/métodos , Células HEK293 , Células HeLa , Humanos , Imãs/química , Nanosferas/ultraestrutura , Neoplasias/sangue , Neoplasias/patologiaRESUMO
Circulating tumour cells (CTCs) draw significant attention as a promising biomarker for cancer prognosis, status monitoring, and metastasis diagnosis. However, the concentration of CTCs in peripheral blood is usually extremely low, thereby requiring enrichment followed by isolation of CTCs prior to detection. An immunomagnetic separation is a promising tool for CTCs enrichment. In this study, a cost-effective magnetic separation method, based on streptavidin-biotin complexation, was developed and the effects of magnetic beads' size in CTCs capture were compared. Magnetic nanobeads which were 25â nm in diameter lead to highest capture efficiency (82.2%) compared with 150â nm magnetic beads and 1â µm microbeads. Based on the streptavidin-biotin system, 25â nm magnetic nanobeads could capture model CTCs over 80% efficiency even at concentrations as low as â¼25â cells/mL that may represent the actual level of CTCs in peripheral blood of cancer patients. Furthermore, the isolated cells remained robust and healthy showing insignificant changes in morphology and behaviour when cultured for 24â h immediately after capture and isolation. The magnetic nanobeads based on streptavidin-biotin complexation showed promise for the easy and efficient capture and isolation of healthy CTCs for further diagnosis and analysis.
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Proteínas de Bactérias/química , Biotina/análogos & derivados , Separação Imunomagnética/métodos , Nanopartículas de Magnetita/química , Células Neoplásicas Circulantes , Proteínas de Bactérias/metabolismo , Biotina/química , Biotina/metabolismo , Citometria de Fluxo , Humanos , Células K562 , Células MCF-7 , Tamanho da PartículaRESUMO
Infectious diseases caused by Listeria monocytogenes pose a great threat to public health worldwide. Therefore, a rapid and efficient method for L. monocytogenes detection is needed. In this study, a biotin-exposure-based immunomagnetic separation (IMS) method was developed. That is, biotinylated antibody was first targeted to L. monocytogenes. Then, streptavidin-functionalized magnetic nanoparticles were added and anchored onto L. monocytogenes cells indirectly through the strong noncovalent interaction between streptavidin and biotin. Biotin-exposure-based IMS exhibited an excellent capability to enrich L. monocytogenes. Specifically, more than 90% of L. monocytogenes was captured when the bacterial concentration was lower than 104â¯colony-forming units (CFU)/mL. Importantly, the antibody dosage was reduced by 10 times of that in our previous study, which used antibody direct-conjugated magnetic nanoparticles. Propidium monoazide (PMA) treatment prior to PCR amplification could eliminate the false-positive results from dead bacteria and detected viable L. monocytogenes sensitively and specifically. For viable L.monocytogenes detection, enriched L. monocytogenes was treated with PMA prior to asymmetric PCR amplification. The detection limits of the combined IMS with nucleic acid lateral flow (NALF) biosensor for viable L. monocytogenes detection were 3.5â¯×â¯103â¯CFU/mL in phosphate buffer solution and 3.5â¯×â¯104â¯CFU/g in lettuce samples. The whole assay process of recognizing viable L. monocytogenes was completed within 6â¯h. The proposed biotin-exposure-mediated IMS combined with a disposable NALF biosensor platform posed no health risk to the end user, and possessed potential applications in the rapid screening and identification of foodborne pathogens.
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Técnicas Biossensoriais , Biotina/química , Separação Imunomagnética , Listeria monocytogenes/isolamento & purificação , Ácidos Nucleicos/química , Ouro/química , Nanopartículas Metálicas/químicaRESUMO
Studies regarding circulating tumor cells (CTCs) have great significance for cancer prognosis, treatment monitoring, and metastasis diagnosis. However, due to their extremely low concentration in peripheral blood, isolation and enrichment of CTCs are the key steps for early detection. To this end, targeting the folic acid receptors (FRs) on the CTC surface for capture with folic acid (FA) using bovine serum albumin (BSA)-tether for multibiotin enhancement in combination with streptavidin-coated magnetic nanoparticles (MNPs-SA) was developed for ovarian cancer CTC isolation. The streptavidin-biotin-system-mediated two-step binding strategy was shown to capture CTCs from whole blood efficiently without the need for a pretreatment process. The optimized parameters for this system exhibited an average capture efficiency of 80%, which was 25% higher than that of FA-decorated magnetic nanoparticles based on the one-step CTC separation method. Moreover, the isolated cells remained highly viable and were cultured directly without detachment from the MNPs-SA-biotin-CTC complex. Furthermore, when the system was applied for the isolation and detection of CTCs in ovarian cancer patients' peripheral blood samples, it exhibited an 80% correlation with clinical diagnostic criteria. The results indicated that FA targeting, in combination with BSA-based multibiotin enhancement magnetic nanoparticle separation, is a promising tool for CTC enrichment and detection of early-stage ovarian cancer.
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Ácido Fólico/análise , Contagem de Células , Feminino , Humanos , Células Neoplásicas Circulantes , Neoplasias Ovarianas , EstreptavidinaRESUMO
A novel sandwich strategy was designed to detect Staphylococcus aureus. The strategy is based on an antibacterial agent that captures bacterial cells and a fluorescein-labeled antibody that acts as the signal-output probe. Vancomycin (Van), which exerts a strong antibacterial effect on Gram-positive bacteria, was utilized as a molecular recognition agent to detect pathogenic bacteria. To effectively concentrate S. aureus, we used bovine serum albumin (BSA) as the amplification carrier to modify magnetic beads (MBs), which were then functionalized with Van. To improve the specificity of the method for S. aureus detection, we adopted fluorescein isothiocyanate (FITC)-tagged pig immunoglobulin G (FITC-pig IgG) as the signal probe and the second recognition agent that bound between the Fc fragment of pig IgG and protein A in the surface of S. aureus. To quantify S. aureus, we measured the fluorescence signal by flow cytometry (FCM). The use of multivalent magnetic nanoprobes (Van-BSA-MBs) showed a high concentration efficiency (>98%) at bacterial concentrations of only 33 colony-forming units (CFU)/mL. Furthermore, the sandwich mode (FITC-pig IgG/SA/Van-BSA-MBs) also showed ideal specificity because Van and IgG bound with S. aureus at two distinct sites. The detection limit for S. aureus was 3.3 × 101 CFU/mL and the total detection process could be completed within 120 min. Other Gram-positive bacteria and Gram-negative bacteria, including Listeria monocytogenes, Bacillus cereus, Cronobacter sakazakii, Escherichia coli O157:H7, and Salmonella Enteritidis, negligibly interfered with S. aureus detection. The proposed detection strategy for S. aureus possesses attractive characteristics, such as high sensitivity, simple operation, short testing time, and low cost.
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Staphylococcus aureus , Animais , Escherichia coli O157 , Citometria de Fluxo , Separação Imunomagnética , Infecções Estafilocócicas , Suínos , VancomicinaRESUMO
Because of the lack of early screening strategies, ovarian cancer is the most deadly cause of gynecologic malignancies. This paper describes an effective method for the separation and detection of ovarian cancer cells from female whole blood, using folic acid (FA) conjugated magnetic iron oxide nanoparticles (IO-FA nanoparticles). The IO nanoparticles were synthesized by thermal decomposition and then covalently conjugated with FA. The IO-FA nanoparticles were stably attached to the surface of ovarian cancer cells by coupling to the over-expressed folate receptor (FR), thereby making the cells magnetic. These "magnetic cells" were separated from the complex blood matrix without destruction under a magnetic field. The separation efficiency was as high as 61.3% when the abundance of spiked ovarian cancer SKOV3 cells was as low as 5 × 10(-5)%. We also successfully detected five (5) out of ten (10) metastatic ovarian cancer patients' whole blood. This study suggested the feasibility of early detecting of metastatic ovarian cancer cells, which may potentially improve the ovarian cancers patients' overall survival rate for clinical applications.