RESUMEN
The application of advances in personalized medicine requires the support of in vitro diagnostic techniques aimed at the accurate, fast, sensitive, and precise determination of selected biomarkers. Herein, a novel optical centrifugal microfluidic device is developed for clinical analysis and point-of-care diagnostics. Based on compact disc technology, the integrated biophotonic system enables multiple immunoassays in miniaturized mode. The disposable microfluidic discs are made in cyclic olefin copolymer (COP), containing arrays of immobilized probes. In the developed approach, up to six patient samples can each be tested simultaneously. A portable instrument (<2 kg) controls the assay and the high-sensitive reproducible optical detection in transmission mode. Also, the instrument incorporates specific functionalities for personalized telemedicine. The device (analytical method, disc platform, reader, and software) has been validated to diagnose IgE-mediated drug allergies, such as amoxicillin and penicillin G. The total and specific IgE to ß-lactam antibiotics were determined in human serum from patients (25 µL). The excellent analytical performances (detection limit 0.24 ng/mL, standard deviation 7-20 %) demonstrated that the developed system could have the potential for a broader impact beyond the allergy field, as it applies to other IVD tests.
Asunto(s)
Biomarcadores , Inmunoglobulina E , Humanos , Biomarcadores/sangre , Inmunoglobulina E/sangre , Inmunoensayo/métodos , Dispositivos Laboratorio en un Chip , Límite de Detección , Hipersensibilidad a las Drogas/diagnóstico , Hipersensibilidad a las Drogas/sangre , Técnicas Analíticas Microfluídicas/instrumentación , Amoxicilina/sangreRESUMEN
The suspicion of beta-lactam allergy directly contributes to the prescription of antibiotics that diverge from the guidelines, increasing antimicrobial resistance, one of the biggest threats to global health. In vitro quantification of specific IgE is very useful for monitoring allergy, as it confirms or rules out immediate beta-lactam drug allergy and helps find safe alternative antibiotic stewardship. However, reliable in vitro quantification of specific IgE to beta-lactam antibiotics by immunoassay is challenging because of the difficulty of having selective immunoreagents, mainly beta-lactam antigens, and its low concentration levels in serum. Thus, reliable and sensitive in vitro tests for multiplex detection of allergy to different beta-lactam antibiotics is currently essential for clinical diagnosis. Nevertheless, the lack of standardization of quantitative in vitro methods makes the comparison and interpretation of the results difficult. Here, as proof of concept, we report an improved multiplex microimmunoassay for beta-lactam allergy in vitro testing standardization. The results revealed that homologous calibration allows reliable quantification of specific IgE in human serum at very low concentrations (144 ng L-1). Moreover, the reproducibility of the results increases 2-fold using an internal standard, achieving accurate quantitative information: 93% and 106% recovery for penicillin and amoxicillin, respectively. We simultaneously evaluated the reliability of the improved multiplexed in vitro method in a cohort of 40 human serum samples and achieved excellent agreement (0.99) with a currently used in vitro test.
Asunto(s)
Hipersensibilidad a las Drogas , beta-Lactamas , Antibacterianos , Hipersensibilidad a las Drogas/diagnóstico , Humanos , Técnicas In Vitro , Estándares de Referencia , Reproducibilidad de los ResultadosRESUMEN
ß-lactams (BLCs) are the most widely used antibiotics and consequently the most common cause of drug allergy in the world. The diagnosis of drug allergy is complex and represents a serious challenge that includes a wide variety of methods. In vitro tests are based on the immunological determination of allergen-specific IgE, but the tests in the market lack the required sensitivity and specificity. In addition, the large sample volume, long incubation times, and single-plex configuration have brought their use into question to complement the clinical information. Here, we report a chemiluminescence immunoassay (CLIA) for multiparametric quantification of specific IgE to penicillin G, penicillin V, amoxicillin, and piperacillin, using histone H1 as a carrier. The developed CLIA allowed the determination of BLC-specific IgE below 0.1 IU/mL, thus allowing identification of allergic patients with better sensitivity, using only 25 µL of a sample (serum). The immunoassay was successfully applied in a cohort of 140 human serum samples, showing good sensitivity (64.6%) as well as specificity (100%), which significantly improve the predictive character of existing BLC-allergy in vitro tests.
Asunto(s)
Especificidad de Anticuerpos , Inmunoensayo/métodos , Inmunoglobulina E/análisis , Límite de Detección , Luminiscencia , beta-Lactamas/inmunología , Humanos , Inmunoglobulina E/sangre , Inmunoglobulina E/inmunologíaRESUMEN
Compact multiplexed biosensors systems hold great potential for diagnosis of diseases where the detection of multiple biomarkers is required. Hypersensitivity Immunoglobulin E mediated syndromes are primary immunodeficiency disorders associated with sensitization to allergens. Assessing immunoglobulin E (IgE) sensitization to allergens is an important strategy for allergy diagnosis. Here, we report for the first time a reliable, flexible and cost-effective optoelectrical biosensor system for the simultaneous determination of total and allergen-specific IgE and IgG, antibodies using an immunogold-silver signal amplification method. The biosensor was constructed on a regular digital versatile disc (DVD) to immobilize a panel of 12 allergen extracts or pure proteins in microarray format, as a proof of concept. The multiplexed biosensor showed a limit of detection of 0.26 IU/mL (624 pg/mL) and 14 ng/mL for IgE and IgG antibodies, respectively. The system was successfully applied in a cohort of 127 human serum samples, showing good sensitivity (97.6%) as well as specificity (85.7%), and an excellent area under the curve (AUC) value was found at 0.977 (confidence interval, CI 0.957 to 0.990) as compared and validated with a reference clinical immunofluorescence assay, confirming an excellent correlation between both techniques. The multiplex biosensor system with on-demand panel composition can be used fully autonomously in clinical or mobile laboratory settings without the need for any additional medical equipment, with which could make it suitable for massive allergy screening campaigns to better define sensitization profiles.
Asunto(s)
Técnicas Biosensibles , Hipersensibilidad , Alérgenos , Humanos , Hipersensibilidad/diagnóstico , Inmunoglobulina E , Inmunoglobulina GRESUMEN
OBJECTIVE: This study aimed to investigate, in healthy volunteers, the relationship between the plasma concentrations of the alpha(1)-adrenoceptor antagonist terazosin and its effects on arterial blood pressure after a single oral administration of terazosin 2 mg. M ethods: Twenty-four healthy volunteers participated in this study. Pharmacokinetic and pharmacodynamic modeling were performed subject by subject. First, plasma concentrations were fitted according to a one-compartment model with first-order absorption and monoexponential elimination. Then the maximum drug-induced decrease (E(max)) effect compartment-model was developed to describe the pharmacodynamic relationships between systolic and diastolic blood pressure and plasma concentrations using the pharmacokinetic parameters that were previously estimated. RESULTS: For systolic blood pressure, E(max) was 29.9 +/- 10.6 mmHg. The corresponding value for decrease in diastolic blood pressure was 39.7 +/- 8.6 mmHg. The effects of terazosin on systolic and diastolic blood pressure could be quantified by an inhibitory E(max) effect compartment model. The obtained first-order rate constant values (0.40 +/- 0.006 h(-)(1) for systolic blood pressure and 0.47 +/- 0.012 h(-)(1) for diastolic blood pressure) were consistent with the rapid development of pharmacological effect. EC(50) (concentration of terazosin that induces an effect at 50% of E(max) values) values were similar for systolic (29.9 +/- 4.3 microg/L) and diastolic (28.7 +/- 4.0 microg/L) blood pressure. A decrease in diastolic blood pressure was the most sensitive response after oral administration of a single dose of terazosin. CONCLUSION: The direct haemodynamic effects of terazosin can be characterized by an E(max) effect compartment model.
Asunto(s)
Antagonistas Adrenérgicos alfa/farmacocinética , Presión Sanguínea/efectos de los fármacos , Prazosina/análogos & derivados , Dolor Abdominal/inducido químicamente , Administración Oral , Antagonistas de Receptores Adrenérgicos alfa 1 , Antagonistas Adrenérgicos alfa/administración & dosificación , Antagonistas Adrenérgicos alfa/farmacología , Adulto , Algoritmos , Área Bajo la Curva , Cromatografía Líquida de Alta Presión , Mareo/inducido químicamente , Relación Dosis-Respuesta a Droga , Femenino , Semivida , Cefalea/inducido químicamente , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Masculino , Prazosina/sangre , Prazosina/farmacocinética , Prazosina/farmacología , Taquicardia/inducido químicamente , Factores de TiempoRESUMEN
OBJECTIVE: Ibuprofen arginate is a salt formulation of ibuprofen designed to reach target concentrations rapidly. The primary objective of this study was to compare the 12-h pharmacokinetic profile of S(+)-ibuprofen following administration of single doses of ibuprofen arginate (600 mg) and dexibuprofen (400 mg) in healthy volunteers. METHODS: Twenty-four volunteers were recruited into an open-label, randomised, two-period, single-centre study with crossover design. RESULTS: Both treatments were well tolerated. Ibuprofen arginate and dexibuprofen showed similar bioavailability for S(+)-ibuprofen. Compared with dexibuprofen, ibuprofen arginate demonstrated a 45% higher maximum concentration (C(max)), and a time to peak concentration (T(max)) 2 h sooner. CONCLUSION: Ibuprofen arginate approaches maximum concentrations of S(+)-ibuprofen faster and higher than dexibuprofen.
Asunto(s)
Antiinflamatorios no Esteroideos/farmacocinética , Arginina/farmacocinética , Ibuprofeno/farmacocinética , Administración Oral , Adolescente , Adulto , Algoritmos , Análisis de Varianza , Antiinflamatorios no Esteroideos/administración & dosificación , Antiinflamatorios no Esteroideos/efectos adversos , Área Bajo la Curva , Arginina/sangre , Arginina/química , Disponibilidad Biológica , Cromatografía Líquida de Alta Presión , Estudios Cruzados , Relación Dosis-Respuesta a Droga , Combinación de Medicamentos , Epistaxis/inducido químicamente , Femenino , Semivida , Cefalea/inducido químicamente , Humanos , Ibuprofeno/sangre , Ibuprofeno/química , Masculino , Tasa de Depuración Metabólica , Estereoisomerismo , Equivalencia Terapéutica , Factores de TiempoRESUMEN
We developed and validated an accurate, sensitive, precise and rapid HPLC method with UV detection for the determination of sirolimus in blood samples from renal, cardiac and hepatic transplants. This method overcomes most of the problems related to previously published assays using a narrow-bore column with base deactivated C18 reversed phase. Whole blood samples were purified by a combination of a precipitating blood matrix with zinc sulphate and a single step liquid-liquid extraction with acetone and 1-chlorobutane. Calibration curves (range 2.5-150 ng/ml), were linear with coefficients of correlation better than 0.996. The relative standard deviation was determined to be less than 8%. The present method has also been validated by a reference laboratory (St. George's Hospital Medical School, London, UK). More of 300 clinical samples have been analysed with this method.