RESUMEN
Modeling and simulation of the central nervous system provides a tool for understanding and predicting the distribution of small molecules throughout the brain tissue and cerebral spinal fluid (CSF), and these efforts often rely on empirical data to make predictions of distributions to move toward a better mechanistic understanding. A physiologically based pharmacokinetic model presented here incorporates multiple means of drug distribution to assemble a model for understanding potential factors that may determine the distribution of drugs across various regions of the brain, including both intra- and extracellular regions. Two classes of parameters are presented. The first concerns regional gross anatomic variability of the brain; the second concerns estimation of unbound fractions of drugs using know membrane phospholipid heterogeneity derived from regional lipid content. The model was then tested by comparing its outcomes to data from published human pharmacokinetic studies of acetaminophen, morphine, and phenytoin. The alignment of model predictions in the plasma, CSF, and tissue concentrations with the published data from studies of those three drugs suggests that the model can be a template for identifying drug localization in the brain. Clearly, knowledge of differentiated drug distribution in the brain is a requisite step in postulating pharmacodynamic and certain disease mechanisms. SIGNIFICANCE STATEMENT: This study concerns the application of heterogenous lipid distribution in brain tissue to predict regional variations in drug distribution in the brain via a mathematical model, thus expanding upon the current understanding of mechanisms of drug distribution in the central nervous system.
Asunto(s)
Encéfalo , Sistema Nervioso Central , Humanos , Encéfalo/fisiología , Acetaminofén , Modelos Teóricos , Lípidos , Modelos BiológicosRESUMEN
Bridging fundamental approaches to model optimization for pharmacometricians, systems pharmacologists and statisticians is a critical issue. These fields rely primarily on Maximum Likelihood and Extended Least Squares metrics with iterative estimation of parameters. Our research combines adaptive chaos synchronization and grid search to estimate physiological and pharmacological systems with emergent properties by exploring deterministic methods that are more appropriate and have potentially superior performance than classical numerical approaches, which minimize the sum of squares or maximize the likelihood. We illustrate these issues with an established model of cortisol in human with nonlinear dynamics. The model describes cortisol kinetics over time, including its chaotic oscillations, by a delay differential equation. We demonstrate that chaos synchronization helps to avoid the tendency of the gradient-based optimization algorithms to end up in a local minimum. The subsequent analysis illustrates that the hybrid adaptive chaos synchronization for estimation of linear parameters with coarse-to-fine grid search for optimal values of non-linear parameters can be applied iteratively to accurately estimate parameters and effectively track trajectories for a wide class of noisy chaotic systems.
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Farmacología/métodos , Algoritmos , Simulación por Computador , Sistemas de Computación , Humanos , Modelos Estadísticos , Dinámicas no LinealesRESUMEN
Nilotinib is a broad-based tyrosine kinase inhibitor with the highest affinity to inhibit Abelson (c-Abl) and discoidin domain receptors (DDR1/2). Preclinical evidence indicates that Nilotinib reduces the level of brain alpha-synuclein and attenuates inflammation in models of Parkinson's disease (PD). We previously showed that Nilotinib penetrates the blood-brain barrier (BBB) and potentially improves clinical outcomes in individuals with PD and dementia with Lewy bodies (DLB). We performed a physiologically based population pharmacokinetic/pharmacodynamic (popPK/PD) study to determine the effects of Nilotinib in a cohort of 75 PD participants. Participants were randomized (1:1:1:1:1) into five groups (n = 15) and received open-label random single dose (RSD) 150:200:300:400 mg Nilotinib vs placebo. Plasma and cerebrospinal fluid (CSF) were collected at 1, 2, 3, and 4 hours after Nilotinib administration. The results show that Nilotinib enters the brain in a dose-independent manner and 200 mg Nilotinib increases the level of 3,4-Dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), suggesting alteration to dopamine metabolism. Nilotinib significantly reduces plasma total alpha-synuclein and appears to reduce CSF oligomeric: total alpha-synuclein ratio. Furthermore, Nilotinib significantly increases the CSF level of triggering receptors on myeloid cells (TREM)-2, suggesting an anti-inflammatory effect. Taken together, 200 mg Nilotinib appears to be an optimal single dose that concurrently reduces inflammation and engages surrogate disease biomarkers, including dopamine metabolism and alpha-synuclein.
Asunto(s)
Encéfalo/metabolismo , Enfermedad de Parkinson/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/administración & dosificación , Pirimidinas/administración & dosificación , Ácido 3,4-Dihidroxifenilacético/líquido cefalorraquídeo , Ácido 3,4-Dihidroxifenilacético/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/análisis , Encéfalo/efectos de los fármacos , Estudios de Cohortes , Dopamina/sangre , Dopamina/metabolismo , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Drogas en Investigación/administración & dosificación , Drogas en Investigación/análisis , Drogas en Investigación/farmacocinética , Ácido Homovanílico/líquido cefalorraquídeo , Ácido Homovanílico/metabolismo , Humanos , Glicoproteínas de Membrana/líquido cefalorraquídeo , Persona de Mediana Edad , Enfermedad de Parkinson/sangre , Placebos/administración & dosificación , Inhibidores de Proteínas Quinasas/sangre , Inhibidores de Proteínas Quinasas/líquido cefalorraquídeo , Inhibidores de Proteínas Quinasas/farmacocinética , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/metabolismo , Pirimidinas/sangre , Pirimidinas/líquido cefalorraquídeo , Pirimidinas/farmacocinética , Receptores Inmunológicos , alfa-Sinucleína/sangre , alfa-Sinucleína/metabolismoRESUMEN
In mathematical pharmacology, models are constructed to confer a robust method for optimizing treatment. The predictive capability of pharmacological models depends heavily on the ability to track the system and to accurately determine parameters with reference to the sensitivity in projected outcomes. To closely track chaotic systems, one may choose to apply chaos synchronization. An advantageous byproduct of this methodology is the ability to quantify model parameters. In this paper, we illustrate the use of chaos synchronization combined with Nelder-Mead search to estimate parameters of the well-known Kirschner-Panetta model of IL-2 immunotherapy from noisy data. Chaos synchronization with Nelder-Mead search is shown to provide more accurate and reliable estimates than Nelder-Mead search based on an extended least squares (ELS) objective function. Our results underline the strength of this approach to parameter estimation and provide a broader framework of parameter identification for nonlinear models in pharmacology.
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Inmunoterapia , Modelos Inmunológicos , Neoplasias/inmunología , Neoplasias/terapia , Dinámicas no Lineales , HumanosRESUMEN
Formaldehyde is a one-carbon, highly water-soluble aldehyde that is used in certain vaccines to inactivate viruses and to detoxify bacterial toxins. As part of the manufacturing process, some residual formaldehyde can remain behind in vaccines at levels less than or equal to 0.02%. Environmental and occupational exposure, principally by inhalation, is a continuing risk assessment focus for formaldehyde. However, exposure to formaldehyde via vaccine administration is qualitatively and quantitatively different from environmental or occupational settings and calls for a different perspective and approach to risk assessment. As part of a rigorous and ongoing process of evaluating the safety of biological products throughout their lifecycle at the FDA, we performed an assessment of formaldehyde in infant vaccines, in which estimates of the concentrations of formaldehyde in blood and total body water following exposure to formaldehyde-containing vaccines at a single medical visit were compared with endogenous background levels of formaldehyde in a model 2-month-old infant. Formaldehyde levels were estimated using a physiologically-based pharmacokinetic (PBPK) model of formaldehyde disposition following intramuscular (IM) injection. Model results indicated that following a single dose of 200 µg, formaldehyde is essentially completely removed from the site of injection within 30 min. Assuming metabolism at the site of injection only, peak concentrations of formaldehyde in blood/total body water were estimated to be 22 µg/L, which is equivalent to a body burden of 66 µg or <1% of the endogenous level of formaldehyde. Predicted levels in the lymphatics were even lower. Assuming no adverse effects from endogenous formaldehyde, which exists in blood and extravascular water at background concentrations of 0.1 mM, we conclude that residual, exogenously applied formaldehyde continues to be safe following incidental exposures from infant vaccines.
Asunto(s)
Reactivos de Enlaces Cruzados/farmacocinética , Formaldehído/farmacocinética , Modelos Biológicos , Reactivos de Enlaces Cruzados/administración & dosificación , Reactivos de Enlaces Cruzados/efectos adversos , Vacuna contra Difteria, Tétanos y Tos Ferina/química , Formaldehído/administración & dosificación , Formaldehído/efectos adversos , Vacunas contra Haemophilus/química , Vacunas contra Hepatitis B/química , Humanos , Lactante , Inyecciones Intramusculares , Vacuna Antipolio de Virus Inactivados/química , Medición de RiesgoRESUMEN
Toxicity Testing in the 21st Century: A Vision and a Strategy, from the National Research Council Committee on Toxicity Testing and Assessment of Environmental Agents, presents a vision wherein toxicology testing moves from feeding test substances to animals for their lifetimes, and assessing clinical laboratory and histopathological changes, to human tissue studies made suitable by recent technological advances in computational biology, toxicogenomics, and the like. This is to be accomplished by elucidating toxicity pathways complemented by targeted testing. The report focuses on the array of available new concepts and attendant technology that the committee considers relevant to its proffer, but, in the final analysis, it describes little in the way of robust strategy for achieving the stated goals. From that perspective, the vision, as described, is no more innovative or far-reaching than goals directed at the utility of cellular metabolism measurements put forth fifty years ago. The report generally lacks the coherence and organization that could have given greater credibility to the committee's deliberative effort.
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Alternativas a las Pruebas en Animales/métodos , Guías como Asunto , Pruebas de Toxicidad/métodos , Toxicología/métodos , Alternativas a las Pruebas en Animales/tendencias , Animales , Predicción , Historia del Siglo XXI , Humanos , National Academy of Sciences, U.S. , Técnicas de Cultivo de Órganos , Pruebas de Toxicidad/tendencias , Toxicología/normas , Estados UnidosRESUMEN
Recent observations suggest the presence of 20S proteasomes (20S) in the lung epithelial lining fluid. However, the physiological relevance of 20S in the alveolar space and possible contribution to disease processes are unknown. Thus, we evaluated whether extracellular proteasomes could have a pathophysiological role in the injured lung using a rat model of lung contusion (LC). Bronchoalveolar lavage fluids (BALF) were obtained at various time points for up to 168 h after LC or sham procedure. Enzyme activities, ELISA and Western blots indicated enzymatically active 20S, the 19S subunit Rpt5 and ubiquitin in BALF. 20S and ubiquitin increased significantly after LC, peaked at 24 h and normalized within 168 h. Mg(2+)/ATP-dependent peptidase activities were detectable 6-24 h after LC. BALF after LC also contained ubiquitin-protein-ligase activity. Addition of Mg(2+)/ATP to BALF after LC led to significant proteolysis and could be prevented with epoxomicin and EDTA. These data suggest for the first time that the Mg(2+)/ATP-dependent 26S proteasome complex exists outside the cell, is released into the lung epithelial lining fluid after LC and contributes to the proteolysis of the bulk of protein in the alveolar space of the injured lung. We infer that proteasome complexes may have a pathophysiological role during lung edema clearance.
Asunto(s)
Bronquios/enzimología , Contusiones/enzimología , Lesión Pulmonar/enzimología , Complejo de la Endopetidasa Proteasomal/metabolismo , Alveolos Pulmonares/enzimología , Adenosina Trifosfato/metabolismo , Animales , Bronquios/patología , Líquido del Lavado Bronquioalveolar/química , Contusiones/patología , Modelos Animales de Enfermedad , Lesión Pulmonar/patología , Magnesio/metabolismo , Masculino , Proteínas/metabolismo , Alveolos Pulmonares/patología , Ratas , Ratas Sprague-Dawley , Factores de Tiempo , Ubiquitina/metabolismoRESUMEN
BACKGROUND: Celiac Disease (CD) is present in 1-16.4% of patients with type 1 diabetes mellitus. The most important serological markers of CD are anti-endomysial (EMA), anti-tissue transglutaminase (tTGA) and antigliadin antibodies (AGA). AIM/HYPOTHESIS: The objective of this work is to determine the frequency of tTGA and/or AGA in latent autoimmune diabetes of adult (LADA) and subjects with type 2 diabetes (T2DM), as well as to evaluate their relation with several clinical and biochemical characteristics. SUBJECTS AND METHODS: Forty three subjects with LADA and 99 with T2DM were studied. The presence of AGA, tTGA was determined in the sera of these patients. The variables: sex, age, duration of diabetes, treatment, body mass index (BMI) and fasting blood glucose concentration were also recorded. RESULTS: No differences were found in the frequency of celiac disease associated antibodies between LADA and T2DM subjects. The presence of celiac disease related antibodies was more frequent in patients with a normal or low BMI. CONCLUSIONS: Celiac disease does not seem to be related with pancreatic autoimmunity in type 2 diabetes. Celiac disease causes a decrease of body mass index in type 2 diabetes while pancreatic islet autoimmunity in this entity masks this effect.
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Autoanticuerpos/inmunología , Enfermedad Celíaca/inmunología , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 2/inmunología , Adulto , Anciano , Índice de Masa Corporal , Femenino , Gliadina/inmunología , Humanos , Masculino , Persona de Mediana Edad , Transglutaminasas/inmunologíaRESUMEN
Coeliac disease (CD) is described as an autoimmune enteropathy associated with the presence of IgG and IgA antigliadin and antitransglutaminase autoantibodies. While of diagnostic significance, the role of these autoantibodies in the immunopathogenesis of CD is elucidated. An inappropriate T cell immune response to gluten is also involved in the pathogenesis of CD, as evidenced by autoantibody switching. The N-glycans released from serum IgG of CD patients and three groups of healthy controls, of differing age ranges, were analysed by NH2-high performance liquid chromatography (HPLC). The fucosylated biantennary N- glycans were the most abundant neutral oligosaccharides; in particular, the agalacto form (G0F) showed a mean value of 42% (s.d. +/- 7.4), 30% (s.d. +/- 5.9), 26% (s.d. +/- 4.2) and 35% (s.d. +/- 6.8) for CD patients, healthy children, healthy adults under 40 and healthy adults over 40 years old, respectively. The ratio of asialo agalacto fucosylated biantenna to asialo monogalacto fucosylated biantenna (G0F)/(G1F) for CD patients showed a significant increase compared to healthy children (P < 0.0002), healthy adults under 40 (P < 0.0002) and healthy adults over 40 years old (P < 0.01). Hypogalactosylation was more pronounced for CD patients than for the patients with other autoimmune diseases such as rheumatoid arthritis or psoriatic arthritis.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Enfermedad Celíaca/inmunología , Inmunoglobulina G/metabolismo , Polisacáridos/análisis , Adulto , Autoanticuerpos/análisis , Estudios de Casos y Controles , Niño , Preescolar , Cromatografía Líquida de Alta Presión/métodos , Gliadina/inmunología , Humanos , Lactante , Oligosacáridos/análisis , Transglutaminasas/inmunologíaRESUMEN
Decompression sickness is a complex phenomenon involving gas exchange, bubble dynamics and tissue response. Relatively simple deterministic compartmental models using empirically derived parameters have been the mainstay of the practice for preventing decompression sickness since the early 1900s. Decades of research have improved our understanding of decompression physiology, and the insights incorporated in decompression models have allowed people to dive deeper into the ocean. However, these efforts have not yet, and are unlikely in the near future, to result in a 'universal' deterministic model that can predict when decompression sickness will occur. Divers using current recreational dive computers need to be aware of their limitations. Probabilistic models based on the estimation of parameters using modern statistical methods from large databases of dives offer a new approach and can provide a means of standardisation of deterministic models. Future improvements in decompression practice will depend on continued improvement in understanding the kinetics and dynamics of gas exchange, bubble evolution and tissue response, and the incorporation of this knowledge in risk models whose parameters can be estimated from large databases of human and animal data.
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Enfermedad de Descompresión/prevención & control , Buceo/efectos adversos , Modelos Biológicos , Intercambio Gaseoso Pulmonar/fisiología , Enfermedad de Descompresión/fisiopatología , HumanosRESUMEN
Monoclonal antibodies directed against recombinant apolipoprotein (a) (r-apo(a)) lacking plasminogen-like KIV-2 repeats were used to identify structurally related conformational epitopes in various members of the plasminogen-prothrombin gene family. A number of procedures including a fibrin-binding inhibition immunoassay and surface plasmon resonance studies were used. Two antibodies (A10.1 and A10.4) recognised common conformational structures in r-apo(a), prothrombin, factor XII, plasminogen and its tissue-type and urokinase-type activators. In contrast, two other antibodies recognised specifically an epitope comprising residues of the lysine-binding site (A10.2) or close to it (A10.5) and inhibited the fibrin-binding function of r-apo(a) (IC(50)=36 pmol/l and 9.76 nmol/l, respectively). Interestingly, these antibodies distinctly recognised the elastase-derived fragments of plasminogen K4 (A10.2) and K1+2+3 (A10.5) without affecting plasminogen binding to fibrin. These results suggest that highly conserved conformational regions are common to various proteins of the plasminogen-prothrombin gene family and are in agreement with the concept that these proteins constitute a monophyletic group derived from an ancestral gene.
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Apolipoproteínas A/química , Fibrina/química , Kringles , Anticuerpos Monoclonales/inmunología , Afinidad de Anticuerpos , Apolipoproteínas A/genética , Apolipoproteínas A/inmunología , Sitios de Unión , Técnicas Biosensibles , Reacciones Cruzadas , Inmunoensayo , Plasminógeno/química , Plasminógeno/genética , Plasminógeno/inmunología , Conformación Proteica , Protrombina/química , Protrombina/genética , Protrombina/inmunología , Proteínas Recombinantes/inmunología , Resonancia por Plasmón de SuperficieAsunto(s)
Apolipoproteínas B/sangre , Hiperlipoproteinemia Tipo II/sangre , Lipoproteína(a)/sangre , Triglicéridos/sangre , Angina de Pecho/sangre , Apolipoproteína A-I/sangre , Arteriosclerosis/sangre , Colesterol/sangre , HDL-Colesterol/sangre , LDL-Colesterol/sangre , Humanos , Persona de Mediana Edad , Infarto del Miocardio/sangre , Valores de ReferenciaRESUMEN
The serine-proteinase domain in human apolipoprotein(a) [apo(a)] and plasminogen exhibit 89% sequence identity including the catalytic triad. Cleavage of the Arg(561)-Val(562) activation site in plasminogen by either tissue- or urokinase-type plasminogen activator results in formation of the fibrinolytic enzyme plasmin. Apo(a) does not contain measurable amidolytic activity nor can it be activated by plasminogen activators. It has been suggested that the latter finding might be explained by the substitution of the plasminogen Arg-Val activation site by Ser-Ile in apo(a). To investigate if introduction of the Arg-Val activation site in apo(a) might result in sensitivity towards plasminogen activators, we expressed wild-type and Arg-Val mutant recombinant apo(a) [r-apo(a)] in human embryonic kidney and hepatocyte cell lines. Free r-apo(a) and lipoprotein-like particles [r-Lp(a)] were obtained in the culture supernatants of transfected 293 and HepG2 cells, respectively. Incubation of mutant r-apo(a)/r-Lp(a) with plasminogen activators produced neither plasmin-like activity nor cleavage at the Arg-Val activation site, even in the presence of various stimulators of plasminogen activation. Our data suggest that the high selectivity of activators for plasminogen activation requires interactions with regions in plasminogen distant from the activation disulfide loop which are not present in apo(a).
Asunto(s)
Apolipoproteínas A/química , Activadores Plasminogénicos/química , Secuencia de Aminoácidos , Apolipoproteínas A/genética , Sitios de Unión , ADN Complementario/química , Activación Enzimática , Humanos , Immunoblotting , Kringles , Lipoproteína(a)/química , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Plásmidos , Plasminógeno/química , Proteínas Recombinantes/química , Serina Endopeptidasas/químicaRESUMEN
BACKGROUND: To date, the most commonly accepted techniques for the screening of coeliac disease are indirect immunofluorescence and enzyme-linked immunosorbent assay (ELISA), which reveal antiendomysium and antigliadin antibodies respectively. We report the use of a simple visual system for coeliac disease screening based on the use of Staphylococcus aureus protein A, which binds to both IgG and IgA, thus avoiding the need for two parallel immunoassays. MATERIALS AND METHODS: Opaque polystyrene microwell strips coated with a wheat gliadin extract were incubated with sera followed by incubation with protein A-colloidal gold conjugate. The resulting colour was compared with that of positive and negative control sera. The procedure took less than an hour. RESULTS: One hundred and forty-five biopsy-proven sera, 94 from active coeliac patients and 51 from non-coeliac patients with diverse gastrointestinal pathologies or diabetes mellitus, were assayed. Ninety of the 94 sera from the active coeliac patients were positive, whereas only 3 of the 51 non-coeliac control subjects were positive. The technique has a sensitivity of 95.7% and a specificity of 94.1%. CONCLUSIONS: The sensitivity and specificity of the visual system are greater than those of most ELISA systems and are similar to those observed with IgA antiendomysium antibodies when tested in the same population. Moreover, it is inexpensive, quick, simple to perform and easy to interpret, i.e. it requires no qualified personnel. It is for these features, together with the outstanding sensitivity and specificity, that we propose this immunoassay as a new test for reliable coeliac disease screening.
Asunto(s)
Enfermedad Celíaca/diagnóstico , Inmunoensayo/métodos , Adolescente , Autoanticuerpos/análisis , Estudios de Casos y Controles , Enfermedad Celíaca/inmunología , Niño , Preescolar , Ensayo de Inmunoadsorción Enzimática , Gliadina/inmunología , Humanos , Inmunoensayo/estadística & datos numéricos , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Fibras Musculares Esqueléticas/inmunología , Sensibilidad y Especificidad , Proteína Estafilocócica ARESUMEN
A cocktail sandwich ELISA based on the employ of two monoclonal antibodies (MAbs) as coating antibodies and a third MAb conjugated to horseradish peroxidase has been developed for the analysis of gluten in foods. Given that each MAb displays a wide specificity spectrum for wheat, barley, rye and oats prolamins, their combination for ELISA ensures a high crossreactivity with most of the potentially toxic gliadin, hordein, secalin and avenin protein family. One of the unprecedented features of the cocktail sandwich ELISA is that it permits for the first time analysis of barley hordeins in foods, which is unattainable using conventional or commercial ELISA kits. Besides, gliadins, hordeins and secalins are recognised to the same extent. The system provides a high detection sensitivity for gliadins, hordeins, secalins and avenins (1.5, 0.05, 0.15 and 12 ng/ml, respectively). The working linear range comprises 3-100 ng/ml with a gliadin detection limit of 1.5 ppm. This limit of detection is even better than that demanded in the latest Codex recommendation, 10 ppm. Cocktail ELISA data were contrasted with those of commercial ELISA kits and confirmed by mass spectrometry, a non-immunological technique which provides evidence for the occurrence of false positive results with the commercial kits.
Asunto(s)
Ensayo de Inmunoadsorción Enzimática/métodos , Glútenes/análisis , Anticuerpos Monoclonales/inmunología , Glútenes/inmunología , Hordeum , Secale , TriticumRESUMEN
The gluten toxic fractions responsible for the mucosal damage in coeliac disease (CD), so-called gliadins, hordeins, secalins and avenins from a large number (30-40) of wheat, barley, rye and oats cultivars respectively, have been mass analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Gliadin, secalin and avenin characteristic mass profiles are nearly identical amongst distinct cultivars from the corresponding cereal, while hordeins profiles show more variability depending on the particular barley cultivar. On the basis of these four distinguishable characteristic mass patterns spreading within the 20,000-40,000 Da range, MALDI-TOF-MS has permitted the direct and simultaneous visualization of gliadins, hordeins, secalins and avenins in foods elaborated with cereal mixtures of wheat, barley, rye and oats. This capacity has been demonstrated by mass analyzing foods made with these four cereals in varying ratios. Thus MALDI-TOF-MS can be preliminarily established as a unique system with the ability to discriminate the specific type of gluten toxic fractions present in food samples.
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Grano Comestible/química , Análisis de los Alimentos/métodos , Glútenes/análisis , Espectrometría de Masas/métodos , Avena/química , Gliadina/análisis , Hordeum/química , Rayos Láser , Proteínas de Plantas/análisis , Prolaminas , Secale/química , Triticum/químicaRESUMEN
Lipoprotein(a) (Lp(a)) is one of the most important independent risk factors for the prediction of premature atherosclerosis. Lp(a) is a low-density lipoprotein (LDL)-like particle which contains a glycoprotein (apoprotein(a)) disulfide linked to apo B-100. We describe a sandwich ELISA based on an anti-apo(a) monoclonal antibody (MAb) and an anti-apo B MAb for the quantitative determination of Lp(a) in human serum. The assay is sensitive, precise and specific. Samples with different apo(a) isoforms had a linear response in a range of 3-70 mg/dl of Lp(a). Correlations between the ELISA and a commercial ELISA, an immunoradiometric assay and electroimmunodiffusion were 0.92, 0.96 and 0.98, respectively. The frequency distribution of Lp(a) concentration in blood donors showed the skew toward the right reported in other populations. Patients with angiographically assessed coronary atherosclerosis had three times higher levels of Lp(a) than those with no signs of coronary atherosclerosis.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Apolipoproteínas A/inmunología , Apolipoproteínas B/inmunología , Ensayo de Inmunoadsorción Enzimática/métodos , Lipoproteína(a)/sangre , Animales , Apolipoproteína B-100 , Donantes de Sangre , Enfermedad de la Arteria Coronaria/sangre , Humanos , Modelos Lineales , Lipoproteína(a)/inmunología , Ratones , Ratones Endogámicos BALB C , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
Rotavirus (RV) is known to be the most common cause of severe diarrhoea in infants and young children, each year leading to an estimated 800,000-900,000 deaths. RV also infects bovines and other species, with high morbidity and mortality. A rapid and simple 'naked-eye' dipstick system was developed to detect human RV in faeces, using nitrocellulose as solid phase, two monoclonal antibodies, and colloidal gold as marker. The system detects 10(4) viral particles (1-2 ng)/g of faeces. For human RV the specificity and sensitivity were 100% when compared with a commercial latex system, and 99% and 98%, respectively, when correlated with traditional RNA-PAGE, and 100% and 98% compared to an ELISA system.
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Heces/microbiología , Rotavirus/aislamiento & purificación , Virología/métodos , Animales , Anticuerpos Monoclonales , Anticuerpos Antivirales , Preescolar , Estudios de Evaluación como Asunto , Gastroenteritis/diagnóstico , Gastroenteritis/virología , Oro Coloide , Humanos , Lactante , Ratones , Rotavirus/inmunología , Infecciones por Rotavirus/diagnóstico , Infecciones por Rotavirus/virología , Sensibilidad y Especificidad , Virología/estadística & datos numéricosRESUMEN
Se desarrolló un sistema ELISA para Apo B empleando el AcM IA/CB-VLDVL.1 como anticuerpo de captura y anticuerpos policlonales de cabra anti Apo B, conjugados con HRP. Este sistema detecta valores superiores de Apo B luego de tratar las muestras con malondialdehído o sulfato cúprico, o luego de la incubación del LDL con células humanas endoteliales en cultivo. El IA/CB-VLDL.1 también reconoce niveles incrementados de Apo B en muestras de suero que han sido incubadas alta temperatura y en presencia de sustancias oxidativas, tales como ázida sódica. Las sustancias antioxidantes como EDTA disminuyen el reconocimiento del determinante antigénico identificado por el AcM IA/CB-VLDL.1. tanto la anbímina humana oxidada, las HDL como la LDL licosiladas no son reconocidas por este AcM. Los niveles de Apo B detectados por el IA/CB-VLDL.1 se correlacionan con los lipoperóxidos en LDL y VLDL alterados con lipoxigenasa en presencia de acidos grasos libres. Este sistema ELISA detectó niveles de Apo B incrementados en muestras de suero de pacientes con aterosclerosis periférica, angiopatía diabética y cardipatía isquémica, con respecto a los sueros de los individuos de distribución de edad y sexo similares, sin manifestaciones clínicas o antecedentes de enfermedad aterosclerótica (AU)