Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 4.356
Filtrar
1.
Sensors (Basel) ; 19(15)2019 Jul 31.
Artículo en Inglés | MEDLINE | ID: mdl-31370234

RESUMEN

This paper proposes a simple approach to optimize the operating frequency band of a lab-on-a-chip based on bio-impedance cytometry for a single cell. It mainly concerns applications in low-conductivity media. Bio-impedance allows for the characterization of low cell concentration or single cells by providing an electrical signature. Thus, it may be necessary to perform impedance measurements up to several tens of megahertz in order to extract the internal cell signature. In the case of single cells, characterization is performed in a very small volume down to 1 pL. At the same time, measured impedances increase from tens of kilo-ohms for physiological liquids up to several mega-ohms for low conductivity media. This is, for example, the case for water analysis. At frequencies above hundreds of kilohertz, parasitic effects, such as coupling capacitances, can prevail over the impedance of the sample and completely short-circuit measurements. To optimize the sensor under these conditions, a complete model of a cytometry device was developed, including parasitic coupling capacitances of the sensor to take into account all the impedances. It appears that it is possible to increase the pass band by optimizing track geometries and placement without changing the sensing area. This assumption was obtained by measuring and comparing electrical properties of yeast cells in a low-conductivity medium (tap water). Decreased coupling capacitance by a factor higher than 10 was obtained compared with a previous non-optimized sensor, which allowed for the impedance measurement of all electrical properties of cells as small as yeast cells in a low-conductivity medium.


Asunto(s)
Técnicas Biosensibles , Recuento de Células/métodos , Dispositivos Laboratorio en un Chip , Saccharomyces cerevisiae/aislamiento & purificación , Capacidad Eléctrica , Conductividad Eléctrica , Impedancia Eléctrica , Citometría de Imagen , Técnicas Analíticas Microfluídicas/métodos , Saccharomyces cerevisiae/fisiología , Análisis de la Célula Individual
2.
Cancer Immunol Immunother ; 68(8): 1341-1350, 2019 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-31324947

RESUMEN

Gastric cancer (GC) is the most common malignant tumor in digestive organs, and the prognosis of GC patients who have undergone surgery remains poor because of frequent recurrence. Therefore, the identification of new markers to predict the outcome of these patients is needed. Monocyte count is a negative prognostic factor associated with inflammation. We investigated the relationship between peripheral monocytes in the peri-operative period and prognosis in GC patients. A high pre-operative monocyte count was identified as a prognostic factor in a retrospective analysis of 278 stage II and III GC patients who underwent curative gastrectomy. In contrast, an increased post-operative monocyte count compared to the pre-operative monocyte count was a marker of poor prognosis, particularly for early relapse. In a prospective analysis of 75 GC patients, a subset of the increased post-operative monocytes was similar to CD14+ HLA-DR- CD11b+ CD33+ cells by flow cytometry, and these monocytes produced IDO and arginase and suppressed T cell functions; therefore, we classified these cells as monocytic myeloid-derived suppressive cells (M-MDSCs). Peri-operative neutrophils and C-reactive protein (CRP), which are also related to inflammation, did not affect the prognosis of GC patients, and a neutrophil immunosuppressive function was not observed. These results suggest that peripheral monocytes in the peri-operative period in GC patients are a useful marker for the prognosis of GC patients, and a subset of increased post-operative monocytes may be characterized as M-MDSCs.


Asunto(s)
Biomarcadores de Tumor , Recuento de Células/métodos , Monocitos/patología , Células Supresoras de Origen Mieloide/patología , Neoplasias Gástricas/diagnóstico , Anciano , Células Cultivadas , Femenino , Citometría de Flujo , Gastrectomía , Humanos , Masculino , Recurrencia Local de Neoplasia , Estadificación de Neoplasias , Periodo Perioperatorio , Pronóstico , Estudios Retrospectivos , Neoplasias Gástricas/inmunología , Neoplasias Gástricas/mortalidad , Análisis de Supervivencia
3.
Anal Chim Acta ; 1077: 216-224, 2019 Oct 24.
Artículo en Inglés | MEDLINE | ID: mdl-31307712

RESUMEN

We designed a smartphone based field-portable cell counter combining the smartphone microscope for bright-field image recording and the smartphone application for automatically cell recognition, counting and analysis. To our best knowledge, it is the first time that a smartphone based cell counter can distinguish and count both live and dead cells simultaneously. Compared to the results obtained by hemocytometer, commercial cell counter and flow cytometer, the proposed device was proved to detect cell concentration and viability accurately within the application range between 105 cells/mL and 107 cells/mL. Though multiple fields of view were measured to increase the sampling amount for error reduction, the whole operations including image recording and processing can still be finished rapidly. Moreover, the proposed device is cost-effective with small size of 170 mm × 113 mm × 168 mm containing a built-in power supply. Considering its advantages as high accuracy, fast speed, low cost, long battery life and compact configuration, it is believed the proposed device is a potential tool applied in on-site cell analysis.


Asunto(s)
Recuento de Células/métodos , Teléfono Inteligente , Animales , Recuento de Células/instrumentación , Diseño de Equipo , Programas Informáticos , Células Vero
4.
Mol Immunol ; 112: 233-239, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-31181422

RESUMEN

BACKGROUND: Regulatory B cells participate in the pathogenesis of autoimmune disease. This study aimed to examine the putative contribution of regulatory B cells to the pathogenesis of DN. The number of circulating CD19+CD24hiCD38hi B cells, CD19+CD24hiCD38hiCD5+ B cells, and CD19+CD24hiCD38hiIL-10+ B cells were significantly lower in DN patients (p < 0.05) than the control group. The number of circulating CD19+CD24hiCD38hi B cells was positively correlated with the levels of eGFR and serum IL-10 levels, but negatively correlated with urinary protein levels in DN patients. Treatment significantly increased the number of CD19+CD24hiCD38hi B cells, CD19+CD24hiCD38hiCD5+ B cells, CD19+CD24hiCD38hiIL-10+ B cells, and the levels of serum IL-10 (p < 0.05). We conclude that regulatory B cells may present new targets for intervention of DN.


Asunto(s)
ADP-Ribosil Ciclasa 1/inmunología , Antígenos CD19/inmunología , Linfocitos B Reguladores/inmunología , Antígeno CD24/inmunología , Nefropatías Diabéticas/inmunología , Glicoproteínas de Membrana/inmunología , Adulto , Anciano , Recuento de Células/métodos , Femenino , Humanos , Interleucina-10/inmunología , Masculino , Persona de Mediana Edad
5.
Psychopharmacology (Berl) ; 236(11): 3329-3339, 2019 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-31201477

RESUMEN

RATIONALE AND OBJECTIVE: The presence of three conspecifics prevents stress-induced decreases in newly proliferated cells and neuroblasts in mouse dentate gyrus (DG). In this study, we sought to determine how many conspecifics are required to exert these protective effects against stress. In addition, we manipulated the physiological status of those conspecifics in the context of their stress-buffering effects and used airborne oxytocin exposure as a substitute for the presence of conspecifics. MATERIALS AND METHODS: Bromodeoxyuridine staining was used to indicate the newly proliferated cells and co-staining with doublecortin to reveal the proliferative neuroblasts. RESULTS: Presentation of three intact and lipopolysaccharide-treated conspecifics prevented the stress-induced decreases in the number of newly proliferated cells and neuroblasts in DG. Presentation of one saline- or oxytocin (OT)-treated conspecific did not exert observable stress-buffering effects. In contrast, airborne oxytocin prevented the stress-induced decreases in DG cell proliferation and early neurogenesis, while pretreatment with L-371,257, a selective OT receptor antagonist, abolished the buffering effects of OT. CONCLUSIONS: Physical interaction with the conspecifics and conspecifics' sickness, at best, play a minor role in mediating the buffering effects against stress-induced decreases in DG cell proliferation or early neurogenesis. Moreover, stress-buffering effects are negligible with the presence of only one conspecific. Finally, airborne OT produced stress-buffering effects possibly via its stimulation of OT receptors. Oxytocin merits further study as a substitute for the stress-buffering effects of companions.


Asunto(s)
Proliferación Celular/fisiología , Células-Madre Neurales/fisiología , Neurogénesis/fisiología , Estrés Psicológico/prevención & control , Animales , Recuento de Células/métodos , Proliferación Celular/efectos de los fármacos , Giro Dentado/citología , Giro Dentado/efectos de los fármacos , Giro Dentado/fisiología , Masculino , Ratones , Ratones Endogámicos BALB C , Células-Madre Neurales/efectos de los fármacos , Neurogénesis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Oxitocina/farmacología , Oxitocina/uso terapéutico , Estrés Psicológico/fisiopatología , Estrés Psicológico/psicología
6.
Biomater Sci ; 7(8): 3359-3372, 2019 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-31231724

RESUMEN

Magnetic nanomaterials have drawn ample attention in the field of biotechnology due to their excellent magnetic properties and biocompatibility. These materials have been widely used for exosome isolation, DNA separation, magnetic resonance imaging, and drug delivery. However, their application in cell isolation has been limited due to the lack of efficient antibody conjugation and instability in aqueous solutions. In this study, we produced hybrid maghemite nanorod/immuno-microgels with high capturing capacity for cell isolation and enumeration. Lepidocrocite (γ-FeOOH) and maghemite (γ-Fe2O3) nanorods with controlled morphology are synthesized using hydrolysis method. The effects of the different synthesis conditions on morphology, phase composition, and magnetic properties of lepidocrocite are studied to determine the best synthesis conditions. We coat the nanorods with chitosan and attach them to the poly(N-isopropylacrylamide-co-acrylic acid) (PNIPAM-AA) microgel through chemical bonding to form a nano/hybrid microstructure. Our results suggest that the hybrid magnetic microgels have more antibody binding capacity and higher cancer cell capturing rate compared to pristine maghemite nanorods. In addition, new cell magnetometery method was applied for cancer cell quantification after capturing step in which different magnetized labelled cells were correlated to the saturation magnetization. In this method, higher concentrations of the primary cell suspension resulted in more binding of the magnetic immuno-microgels to the cells which was shown as saturation magnetization drop in the microgel-cell complex.


Asunto(s)
Recuento de Células/métodos , Separación Celular/métodos , Compuestos Férricos/química , Nanoestructuras/química , Resinas Acrílicas/química , Geles , Nanotubos/química , Propiedades de Superficie
7.
Cell Prolif ; 52(4): e12587, 2019 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-31206838

RESUMEN

OBJECTIVES: Cellular aggregates are readily applicable in cell-based therapy. The effects of agitation and inoculation density on the aggregation of cells in spinner flask and the molecular mechanism of aggregation were investigated. MATERIALS AND METHODS: The aggregation kinetics of cells in spinner flask was evaluated with bovine articular chondrocytes (bACs), rabbit bone marrow-derived mesenchymal stem cells (rMSCs) and their mixture. The morphology of cellular aggregates was studied with scanning electron microscopy and gene expression of cell adhesion-related molecules was analysed. RESULTS: It was shown that suspension culture in spinner flask induced the aggregation of bACs and rMSCs. Both cells exhibited increased aggregation rate and aggregate size with decreasing agitation rate and increasing cell inoculation density. Additionally, aggregate size increased with extended culture time. By analysing gene expression of integrin ß1 and cadherin, it was indicated that these molecules were potentially involved in the aggregation process of bACs and rMSCs, respectively. Aggregates composed of both bACs and rMSCs were also prepared, showing rMSCs in the core and bACs in the periphery. CONCLUSIONS: Cellular aggregates were prepared in dynamic suspension culture using spinner flask, the key parameters to the aggregation process were identified, and the molecular mechanism of aggregation was revealed. This would lay a solid foundation for the large-scale production of cellular aggregates for cell-based therapy, such as cartilage regeneration.


Asunto(s)
Condrocitos/citología , Células Madre Mesenquimatosas/citología , Animales , Médula Ósea/metabolismo , Médula Ósea/fisiología , Bovinos , Adhesión Celular/fisiología , Moléculas de Adhesión Celular/metabolismo , Recuento de Células/métodos , Técnicas de Cultivo de Célula/métodos , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Células Cultivadas , Condrocitos/metabolismo , Expresión Génica/fisiología , Células Madre Mesenquimatosas/metabolismo , Conejos
8.
Sensors (Basel) ; 19(12)2019 Jun 20.
Artículo en Inglés | MEDLINE | ID: mdl-31226728

RESUMEN

BACKGROUND: Biomass measurement and monitoring is a challenge in a number of biotechnology processes where fast, inexpensive, and non-contact measurement techniques would be of great benefit. Magnetic induction spectroscopy (MIS) is a novel non-destructive and contactless impedance measurement technique with many potential industrial and biomedical applications. The aim of this paper is to use computer modeling and experimental measurements to prove the suitability of the MIS system developed at the University of South Wales for controlled biomass measurements. METHODS: The paper reports experimental measurements conducted on saline solutions and yeast suspensions at different concentrations to test the detection performance of the MIS system. The commercial electromagnetic simulation software CST was used to simulate the measurement outcomes with saline solutions and compare them with those of the actual measurements. We adopted two different ways for yeast suspension preparation to assess the system's sensitivity and accuracy. RESULTS: For saline solutions, the simulation results agree well with the measurement results, and the MIS system was able to distinguish saline solutions at different concentrations even in the small range of 0-1.6 g/L. For yeast suspensions, regardless of the preparation method, the MIS system can reliably distinguish yeast suspensions with lower concentrations 0-20 g/L. The conductivity spectrum of yeast suspensions present excellent separability between different concentrations and dielectric dispersion property at concentrations higher than 100 g/L. CONCLUSIONS: The South Wales MIS system can achieve controlled yeast measurements with high sensitivity and stability, and it shows promising potential applications, with further development, for cell biology research where contactless monitoring of cellular density is of relevance.


Asunto(s)
Biomasa , Biotecnología , Recuento de Células/métodos , Levaduras/crecimiento & desarrollo , Simulación por Computador , Espectroscopía Dieléctrica , Campos Electromagnéticos , Solución Salina/química
9.
J Nanobiotechnology ; 17(1): 59, 2019 May 04.
Artículo en Inglés | MEDLINE | ID: mdl-31054582

RESUMEN

BACKGROUND: Technology enabling the separation of rare circulating tumor cells (CTCs) provides the potential to enhance our knowledge of cancer metastasis and improve the care of cancer patients. Modern detection approaches commonly depend on tumor antigens or the physical size of CTCs. However, few studies report the detection of CTCs by the electrical differences between cancer cells and normal cells. RESULTS: In this study, we report a procedure for capturing CTCs from blood samples using electrically charged superparamagnetic nanoparticles (NPs). We found that only positively charged NPs attached to cancer cells, while negatively charged NPs did not. The capture method with positively charged NPs offered a sensitivity of down to 4 CTCs in 1 mL blood samples and achieved a superior capture yield (> 70%) for a high number of CTCs in blood samples (103-106 cells/mL). Following an in vitro evaluation, S180-bearing mice were employed as an in vivo model to assess the specificity and sensitivity of the capture procedure. The number of CTCs in blood from tumor-bearing mice was significantly higher than that in blood from healthy controls (on average, 75.8 ± 16.4 vs. zero CTCs/100 µL of blood, p < 0.0001), suggesting the high sensitivity and specificity of our method. CONCLUSIONS: Positively charged NPs combined with an in vivo tumor model demonstrated that CTCs can be distinguished and isolated from other blood cells based on their electrical properties.


Asunto(s)
Neoplasias de la Mama/diagnóstico , Óxido Ferrosoférrico/química , Nanopartículas de Magnetita/química , Células Neoplásicas Circulantes/metabolismo , Adulto , Anciano , Animales , Antígenos de Neoplasias/metabolismo , Técnicas Biosensibles/métodos , Sangre , Recuento de Células/métodos , Línea Celular Tumoral , Separación Celular/métodos , Femenino , Humanos , Magnetismo , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Modelos Animales , Tamaño de la Partícula , Sensibilidad y Especificidad , Propiedades de Superficie
10.
Biosens Bioelectron ; 137: 222-228, 2019 Jul 15.
Artículo en Inglés | MEDLINE | ID: mdl-31121459

RESUMEN

Electrochemically active bacteria (EAB) use extracellular electron transfer (EET) to exchange electron with extracellular acceptors. Previous studies regarding the measurement of EAB were based on either extracellular reduction or oxidation. In this work, we developed a simple electrochemiluminescence (ECL) assay for the identification and detection of EAB. The results of this proposed method revealed that EET of EAB influenced the content of dissolved oxygen and the formation of Ru(bpy)32+• thus leading to qualitative changes of the ECL signal. EAB with the ability of extracellular reduction (such as Shewanella oneidensis MR-1) gave enhanced signal on ECL emission while those displaying the ability of extracellular oxidation (i.e., Sulfobacillus acidophilus) showed the opposite effect on ECL emission, but non-EAB (i.e., Escherichia coli) did not. These changes in ECL intensity were also proportional to the cell density that could be quantitatively detected in the concentration range of (1.1 ±â€¯1) × 105-212 ±â€¯2 CFU/mL (i.e. Shewanella oneidensis MR-1). Moreover, the measurement of the ability of EAB using this approach was in agreement with measurements using the dissimilatory Fe(III) reduction method. Compared to previous reports, this method displayed a continual and steady ECL signal that allowed accurate measurements of EAB. Most important, only a low cell density was needed in this Ru(bpy)32+ - based ECL method, which is beneficial for cell detection.


Asunto(s)
Técnicas Biosensibles , Recuento de Células/métodos , Compuestos Férricos/química , Shewanella/aislamiento & purificación , Técnicas Electroquímicas , Transporte de Electrón , Mediciones Luminiscentes , Fotometría , Shewanella/química
11.
Anal Chim Acta ; 1071: 36-43, 2019 Sep 13.
Artículo en Inglés | MEDLINE | ID: mdl-31128753

RESUMEN

This study describes for the first time the development of 3D printed microfluidic devices with integrated electrodes for label-free counting of E. coli cells incorporated inside droplets based on capacitively coupled contactless conductivity detection (C4D). Microfluidic devices were fully fabricated by 3D printing in the T-junction shape containing two channels for disperse and continuous phases and two sensing electrodes for C4D measurements. The disperse phase containing E. coli K12 cells and the continuous phase containing oil and 1% Span® 80 were pumped through flow rates fixed at 5 and 60 µL min-1, respectively. The droplets with incorporated cells were monitored in the C4D system applying a 500-kHz sinusoidal wave with 1 Vpp amplitude. The generated droplets exhibited a spherical shape with average diameter of 321 ±â€¯9 µm and presented volume of 17.3 ±â€¯0.5 nL. The proposed approach demonstrated ability to detect E. coli cells in the concentration range between 86.5 and 8650 CFU droplet-1. The number of cells per droplet was quantified through the plate counting method and revealed a good agreement with the Poisson distribution. The limit of detection achieved for counting E. coli cells was 63.66 CFU droplet-1. The label-free counting method has offered instrumental simplicity, low cost, high sensitivity and compatibility to be integrated on single microfluidic platforms entirely fabricated by 3D printing, thus opening new possibilities of applications in microbiology.


Asunto(s)
Recuento de Células/métodos , Conductividad Eléctrica , Técnicas Electroquímicas/métodos , Escherichia coli K12/aislamiento & purificación , Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas/métodos , Técnicas Electroquímicas/instrumentación , Electrodos , Límite de Detección , Técnicas Analíticas Microfluídicas/instrumentación , Impresión Tridimensional
12.
N Z Vet J ; 67(4): 203-209, 2019 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-31023170

RESUMEN

Aims: To assess the use of different cut-points based on individual cow somatic cell counts (SCC) to define cows with intramammary infection (IMI) at drying-of, in a herd with a high mean bulk tank SCC. Methods: Results for SCC from four herd tests during lactation and bacterial culture of milk samples collected before drying-off were obtained for 139 cows from a herd with an average bulk milk SCC of >300,000 cells/mL over the final 4 months of the 2006/07 lactation. Based on culture results, cows were defined as being infected with a major (Staphylococcus aureus, Streptococcus uberis or Nocardia spp.) or any pathogen. Receiver-operator characteristics (ROC) curves were used to determine optimum cut-points for maximum, average and last herd test SCC, for predicting IMI. Multivariable logistic regression models were used to determine which variables were associated with IMI, and the sensitivity (Se), specificity (Sp) and positive predictive value (PPV) were determined for different cut-points. Results: At the cow level, 75/139 (54.0%) cows had IMI with a major pathogen and 123/139 (88.5%) with any pathogen. A SCC ≥150,000 cells/mL at ≥2 herd tests and a SCC ≥299,000 cells/mL at the last herd test, for cows aged ≥4 years, were associated with IMI with a major pathogen at drying-off (p<0.05). A SCC ≥150,000 cells/mL at ≥2 herd tests was associated with IMI with any pathogen at drying-off (p<0.001). A cut-point of ≥150,000 cells/mL at any herd test had the highest Se (0.97 and 0.94), but the lowest Sp (0.19 and 0.44) and PPV (0.58 and 0.93) for infection with major and any pathogens, respectively. A cut-point of ≥150,000 cells/mL at ≥2 herd tests doubled the Sp and increased the PPV without large decreases in test Se for infection with either a major or any pathogen. Conclusions and clinical relevance: In this herd with a high bulk milk SCC, use of a cut-point of a SCC ≥150,000 cells/mL at any herd test to define IMI would be appropriate, where the goal at drying-off is to ensure that cows infected with any pathogen receive antimicrobial treatment. Where the goal is to reduce the use of antimicrobial dry cow therapy in uninfected cows while limiting the number of infected cows not being treated, use of a cut-point of SCC ≥150,000 cells/mL at ≥2 herd tests to define IMI may be more appropriate.


Asunto(s)
Recuento de Células/métodos , Infecciones por Bacterias Grampositivas/veterinaria , Mastitis Bovina/diagnóstico , Mastitis Bovina/microbiología , Leche/microbiología , Animales , Bovinos , Industria Lechera , Femenino , Bacterias Grampositivas/aislamiento & purificación , Infecciones por Bacterias Grampositivas/diagnóstico , Infecciones por Bacterias Grampositivas/epidemiología , Modelos Logísticos , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/epidemiología , Nueva Zelanda/epidemiología , Curva ROC , Estudios Retrospectivos
13.
Methods Mol Biol ; 1979: 155-176, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31028637

RESUMEN

Peripheral blood mononuclear cells (PBMCs) are blood cells that are a critical part of the immune system used to fight off infection. However, due to the complexity of PBMCs, which contain multiple different cell types, studying the function of the individual cell types can be difficult, and often studies rely on bulk measurements. Here, we describe the analysis of PBMCs using single-cell RNA-sequencing in droplets. Data from these studies allow for the identification and quantification of the subpopulation of cells that make up the PBMC sample. In addition, differential gene expression between cell types and samples can be assessed.


Asunto(s)
Leucocitos Mononucleares/metabolismo , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Recuento de Células/métodos , Separación Celular/métodos , ADN Complementario/genética , Diseño de Equipo , Secuenciación de Nucleótidos de Alto Rendimiento/instrumentación , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Leucocitos Mononucleares/citología , Receptores de Lipopolisacáridos/análisis , Monocitos/citología , Monocitos/metabolismo , Transcripción Reversa , Análisis de Secuencia de ARN/instrumentación , Análisis de la Célula Individual/instrumentación , Flujo de Trabajo
14.
J Dairy Sci ; 102(6): 5419-5429, 2019 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-30954252

RESUMEN

Timely and accurate identification of cows with intramammary infections is essential for optimal udder health management. Various sensor systems have been developed to provide udder health information that can be used as a decision support tool for the farmer. Among these sensors, the DeLaval Online Cell Counter (DeLaval, Tumba, Sweden) provides somatic cell counts from every milking at cow level. Our aim was to describe and evaluate diagnostic sensor properties of these online cell counts (OCC) for detecting an intramammary infection, defined as an episode of subclinical mastitis or a new case of clinical mastitis. The predictive abilities of a single OCC value, rolling averages of OCC values, and an elevated mastitis risk (EMR) variable were compared for their accuracy in identifying cows with episodes of subclinical mastitis or new cases of clinical mastitis. Detection of subclinical mastitis episodes by OCC was performed in 2 separate groups of different mastitis pathogens, Pat 1 and Pat 2, categorized by their known ability to increase somatic cell count. The data for this study were obtained in a field trial conducted in the dairy herd of the Norwegian University of Life Sciences. Altogether, 173 cows were sampled at least once during a 17-mo study period. The total number of quarter milk cultures was 5,330. The most common Pat 1 pathogens were Staphylococcus epidermidis, Staphylococcus aureus, and Streptococcus dysgalactiae. The most common Pat 2 pathogens were Corynebacterium bovis, Staphylococcus chromogenes, and Staphylococcus haemolyticus. The OCC were successfully recorded from 82,182 of 96,542 milkings during the study period. For episodes of subclinical mastitis the rolling 7-d average OCC and the EMR approach performed better than a single OCC value for detection of Pat 1 subclinical mastitis episodes. The EMR approach outperformed the OCC approaches for detection of Pat 2 subclinical mastitis episodes. For the 2 pathogen groups, the sensitivity of detection of subclinical mastitis episodes was 69% (Pat 1) and 31% (Pat 2), respectively, at a predefined specificity of 80% (EMR). All 3 approaches were equally good at detecting new cases of clinical mastitis, with an optimum sensitivity of 80% and specificity of 90% (single OCC value).


Asunto(s)
Infecciones por Corynebacterium/veterinaria , Mastitis Bovina/diagnóstico , Leche/microbiología , Sistemas en Línea , Infecciones Estafilocócicas/veterinaria , Infecciones Estreptocócicas/veterinaria , Animales , Infecciones Asintomáticas , Bovinos , Recuento de Células/métodos , Recuento de Células/veterinaria , Corynebacterium/aislamiento & purificación , Infecciones por Corynebacterium/diagnóstico , Infecciones por Corynebacterium/microbiología , Femenino , Lactancia , Estudios Longitudinales , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/microbiología , Mastitis Bovina/microbiología , Infecciones Estafilocócicas/diagnóstico , Infecciones Estafilocócicas/microbiología , Staphylococcus/aislamiento & purificación , Infecciones Estreptocócicas/diagnóstico , Infecciones Estreptocócicas/microbiología , Streptococcus/aislamiento & purificación
15.
Biosens Bioelectron ; 132: 230-237, 2019 May 01.
Artículo en Inglés | MEDLINE | ID: mdl-30877888

RESUMEN

A novel sensor platform modified with clay-protein based composite nanoparticles (Mt-HSA NCs) was developed to be used in electrochemical cytosensing application for the first time. The nanocomposite synthesized with desolvation method was structurally clarified by various characterization methods. Then, the working electrode was constructed by modifying the surface of the disposable pencil graphite (PGE) with physical adsorption to perform a simple sensor system. The characterization studies proved that the Mt-HSA NCs modified surface had a biocompatible, hydrophilic and large surface area where cancer cells can easily attach to the surface. As a diagnostic method, electrochemical impedance spectroscopy (EIS), which has become very popular in recent years, was carried out. The linearity range was found as from 1.5 × 102 to 7.5 × 106 breast cancer (MCF-7) cells and the limit of detection was calculated as 148 cells mL-1. From these results, a simple, effortless, cost effective and rapid electrochemical impedimetric sensor system for the diagnosis of breast cancer was developed by examining the interaction of Mt-HSA NCs/PGE surface with MCF-7 cells.


Asunto(s)
Bentonita/química , Técnicas Biosensibles/métodos , Neoplasias de la Mama/diagnóstico , Espectroscopía Dieléctrica/métodos , Nanocompuestos/química , Albúmina Sérica Humana/química , Recuento de Células/métodos , Línea Celular , Femenino , Grafito/química , Humanos , Células MCF-7 , Nanocompuestos/ultraestructura , Nanopartículas/química , Nanopartículas/ultraestructura
16.
Prev Vet Med ; 165: 44-51, 2019 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-30851927

RESUMEN

Milking-time testing (MTT) is a method for evaluating the vacuum conditions in the teatcup during milking. The purpose is to evaluate the possible impact of the milking and milking equipment on udder health and milk quality. The method is commonly implemented by herd health advisory services, but results are interpreted empirically due to lack of scientific documentation on relationships between MTT result variables and objective measures of udder health. The current study was conducted to increase our understanding of associations between cow-level differences in composite milk somatic cell count (CMSCC) and MTT results in dairy cows milked in 3 different milking systems; automatic milking systems (AMS), milking parlors, and pipeline milking systems. Data from 7069 cows (predominantly Norwegian Red breed) in 1009 herds were used in a cross-sectional study. Multilevel linear regression models with a random intercept at herd level were used to describe relationships between CMSCC (on logarithmic scale) and the following MTT explanatory variables: average vacuum level in the short milk tube and mouthpiece chamber in the main milking and overmilking periods, the duration of these two periods, and vacuum stability, measured by sudden vacuum drops in the short milk tube. The models were corrected for the herd effect, mastitis history and differences in milk yield, lactation stage and parity between cows. Separate models were run for AMS, milking parlors, and pipeline milking systems, because this approach allowed for comparison between systems and for evaluation of the herd effect independently of milking system. The models described 8-10 % of the variation in CMSCC, indicating that MTT could only explain a relatively small proportion of a large total variation in CMSCC. In most observations, vacuum levels in the short milk tube during main milking were within the range recommended by the International Organization for Standardization. The results from our multivariable models showed decreasing CMSCC with increasing vacuum level in the short milk tube during the main milking period in AMS and milking parlors. Similarly, decreasing CMSCC was also associated with increasing duration of the main milking period in all 3 systems. These relationships are important for the interpretation of MTT results under practical conditions; finding high vacuum levels and long milking durations in a MTT is not associated with elevated CMSCC. In AMS herds, we also found indications that the relationships were different for cows where a case of mastitis had been treated before the MTT.


Asunto(s)
Industria Lechera/métodos , Mastitis Bovina/epidemiología , Leche/citología , Animales , Bovinos , Recuento de Células/métodos , Recuento de Células/veterinaria , Estudios Transversales , Industria Lechera/instrumentación , Femenino , Mastitis Bovina/etiología , Factores de Riesgo , Factores de Tiempo
17.
Blood ; 133(13): 1406-1414, 2019 03 28.
Artículo en Inglés | MEDLINE | ID: mdl-30728141

RESUMEN

Cells and the molecular processes underlying their behavior are highly dynamic. Understanding these dynamic biological processes requires noninvasive continuous quantitative single-cell observations, instead of population-based average or single-cell snapshot analysis. Ideally, single-cell dynamics are measured long-term in vivo; however, despite progress in recent years, technical limitations still prevent such studies. On the other hand, in vitro studies have proven to be useful for answering long-standing questions. Although technically still demanding, long-term single-cell imaging and tracking in vitro have become valuable tools to elucidate dynamic molecular processes and mechanisms, especially in rare and heterogeneous populations. Here, we review how continuous quantitative single-cell imaging of hematopoietic cells has been used to solve decades-long controversies. Because aberrant cell fate decisions are at the heart of tissue degeneration and disease, we argue that studying their molecular dynamics using quantitative single-cell imaging will also improve our understanding of these processes and lead to new strategies for therapies.


Asunto(s)
Hematopoyesis , Análisis de la Célula Individual/métodos , Animales , Recuento de Células/métodos , Rastreo Celular/métodos , Biología Computacional/métodos , Células Madre Hematopoyéticas/citología , Humanos , Células Mieloides/citología
18.
Biotechniques ; 66(2): 99-102, 2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30744408

RESUMEN

Cellular proliferation and migration are crucial during development, regeneration and disease. Methods to quantify these processes are available; however, many are time consuming and require specialized equipment and costly reagents. Simple cell counts (proliferation analysis) and the scratch assay (migration analysis) are favorable methods due to their simplicity and cost-effectiveness; however, they rely on subjective and labor-intensive manual analysis, resulting in low throughput. We have developed optimized protocols to rapidly and accurately quantify adherent cell number and wound area using ImageJ, an open-source image processing program. Notably, these adaptable protocols facilitate quantification with significantly greater accuracy than manual identification.


Asunto(s)
Recuento de Células/métodos , Movimiento Celular/genética , Proliferación Celular/genética , Imagen Molecular/métodos , Humanos , Procesamiento de Imagen Asistida por Computador
19.
Clin Chem ; 65(4): 549-558, 2019 04.
Artículo en Inglés | MEDLINE | ID: mdl-30737205

RESUMEN

INTRODUCTION: Circulating tumor cells (CTCs) may be used to improve cancer diagnosis, prognosis, and treatment. However, because knowledge regarding CTC biology is limited and the numbers of CTCs and CTC-positive cancer patients are low, progress in this field is slow. We addressed this limitation by combining diagnostic leukapheresis (DLA) and microfluidic enrichment to obtain large numbers of viable CTCs from metastasized breast cancer patients. METHODS: DLA was applied to 9 patients, and 7.5 mL of peripheral blood was drawn. CTCs were enriched with the Parsortix™ system. The quality of CTCs from fresh and cryopreserved DLA products was tested, and CTCs were cultured in vitro. Single uncultured and cultured CTCs were isolated by micromanipulation to determine different parameters, such as genomic aberrations and mutation profiles of selected tumor-associated genes. Expression levels of estrogen receptor and HER2/neu were monitored during in vitro culture. RESULTS: Viable CTCs from peripheral blood and fresh or frozen DLA products could be enriched. DLA increased the likelihood of successful CTC culture. Cryopreserved DLA products could be stored with minimal CTC loss and no overt reduction in the tumor cell quality and viability during an observation period of up to 3 years. The analyzed parameters did not change during in vitro culture. DLA samples with high CTC numbers and lower ratios of apoptotic CTCs were more likely to grow in culture. CONCLUSIONS: The increased CTC numbers from fresh or cryopreserved DLA products facilitate multiple functional and molecular analyses and, thus, could improve our knowledge of their biology.


Asunto(s)
Biomarcadores de Tumor/sangre , Neoplasias de la Mama/sangre , Leucaféresis/métodos , Células Neoplásicas Circulantes , Neoplasias de la Mama/patología , Recuento de Células/métodos , Humanos , Microfluídica/métodos , Células Neoplásicas Circulantes/patología
20.
Int J Lab Hematol ; 41(2): 277-286, 2019 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-30758900

RESUMEN

INTRODUCTION: Cellular analysis in body fluids (BFs) provides important diagnostic information in various pathological settings. This study was hence aimed at comparing automated cell count obtained with Mindray BC-6800 (BC-BF) vs Sysmex XN-series (XN-BF) and evaluating other quantitative and qualitative information provided by these analyzers in ascitic (AF), pleural (PF), synovial (SF), and cerebrospinal (CSF) fluids. METHODS: Three hundred and fifty-one samples (99 AFs, 45 PFs, 75 SFs, and 132 CSFs) were analyzed in parallel with BC-BF, XN-BF, and optical microscopy (OM). The study also included the assessment of diagnostic agreement among BC-BF, XN-BF, and OM. RESULTS: The comparison of BC-BF vs XN-BF yielded slopes of Passing and Bablok regression always comprised between 0.9 and 1.0 except for EO-BF and HF-BF, whilst the intercepts ranged from -0.4 for MN-BF and 12.0 for PMN-BF. The bias was comprised between -103.3% and 67.1% for HF-BF and EO-BF, respectively. A significant bias was found for TC-BF, WBC-BF, HF-BF (negative bias) and for PMN-BF and EO-BF (positive bias). The agreement (Cohen's kappa) between XN-BF and BC-BF was always ≥0.7, ranging between 0.87 in CSFs and 0.94 in AFs, and that with OM was similar (ie, 0.85 and 0.96). CONCLUSION: The cytometric analysis of BF samples using BC-BF and XN-BF is clinically favorable when appropriately combined with OM. Quantitative and qualitative parameters displayed optimal agreement, whilst instrument-specific cut-offs should be defined and implemented for HF-BF and EO-BF. Further efforts should be made for achieving better harmonization in cytometric analysis of BF samples.


Asunto(s)
Líquido Ascítico/citología , Líquido Cefalorraquídeo/citología , Citometría de Flujo , Líquido Sinovial/citología , Recuento de Células/instrumentación , Recuento de Células/métodos , Femenino , Citometría de Flujo/instrumentación , Citometría de Flujo/métodos , Humanos , Masculino
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA