Selective in situ functionalization of biosensors on LOC devices using laminar co-flow.

Many applications involving lab-on-a-chip (LOC) devices are prevented from entering the market because of difficulties to achieve mass production and impart suitable properties allowing long-term storage. To integrate biosensors on these microfluidic chips, one of the main restrictions is the fabrication and stability of the molecular modifications that must be performed on the surfaces of the sensors for a given application. The complexity of the problem increases exponentially when the LOC integrates several of these sensors. Here we present a system based on laminar co-flow to perform an on-chip selective surface bio-functionalization of LOC-integrated sensors. This method has the advantage that the surface modification protocols are performed in situ before analyte detection. This approach reduces the burdens during LOC fabrication, keeping the required reagents stored outside of the detection structure in suitable wet conditions. The proof of concept is demonstrated through an optical characterization followed by electronic detection based on a novel differential impedance measurement setup. The system can be easily scaled to incorporate several sensors with distinct biosensing targets in a single chip.

Validation of ATP luminometry for rapid and accurate titration of Mycoplasma hyopneumoniae in Friis medium and a comparison with the color changing units assay.

Limited reports are available on the growth response of Mycoplasma hyopneumoniae in Friis medium and the routinely used color changing units (CCU) assay has not yet been profoundly compared with other titration methods. Firstly, growth kinetics of 7 diverse M. hyopneumoniae isolates were followed by ATP luminometry in five Friis medium batches. Secondly, results of the CCU and ATP assays were compared hereby evaluating the methods. Growth curves of all isolates had log, stationary and senescence phases, and reached similar maximal titres when cultured in the same batch of Friis medium. Doubling times (Tds) of the isolates grown in slowly shaken cultures varied between 4.8 and 7.8 h. Maximal titres, Tds, growth phase in which the phenol red indicator turned from red to yellow due to acidification by mycoplasmal metabolism, and the length of the stationary phase varied depending on the Friis medium batch. The effect of static vs. shaking culture conditions on the Td depended on the isolate. ATP and CCU assays obtained similar growth curves, but when maximal levels were reached the CCU titre dropped earlier than the ATP titre. During log phase, CCU and ATP titres were strongly linearly linked. We developed a model enabling transformation of ATP into CCU titres or vice versa. The calculated amount of ATP per CCU (1.77 amol ATP/ml) indicated that the CCU assay likely underestimates the actual cell concentration. When titres were determined as means of 3 measurements, the ATP assay was 7 times more accurate and had 11-fold lower outliers than the CCU assay. Unlike the CCU assay, luminometry only requires one measurement to obtain sufficient accuracy. It was concluded that the ATP assay constitutes a valuable robust alternative for reproducible real-time titre assessment of freshly grown M. hyopneumoniae cultures. It is faster, more accurate and time, work and cost efficient compared to the CCU assay. The assay is preferred to better standardise and describe M. hyopneumoniae cultures used in various experiments.

[The importance of comorbidity in nephrologists' assessment of acute renal failure]. / La importancia de la comorbilidad en el fracaso renal agudo valorado por los nefrólogos.

AIMS: To study the features of acute renal failure (ARF) in our hospital and to determine prognosis and mortality associated factors. METHODS: Retrospective study of ARF episodes during a two years period (2005-2007). ARF was considered when a sudden rise in serum creatinine concentration was more than 0,5 mg/dl in patients with normal renal function and more than 1 mg/dl in patients with previous mild to moderate chronic renal failure. We analyzed epidemiologic, clinical, laboratories results, therapeutics and prognosis factors. RESULTS: Two hundred and one patients were evaluated (62,7% males; Age= 67,35 16,38 years (63,68%>65 years); Comorbility Index of Charlson 3,49 2,43). 115 episodes presented in patients with previous renal failure. ARF was pre-renal in 52,7%, renal in 34,8% and post-renal in 8,5%. 35,8% of ARF patients had oliguria or anuria. The mean duration of ARF/hospitalization was 22,47 days (22,47 21,83). The percentage of resolved ARF was 70,1%. Mortality was 30,8%. The univariated analysis showed comorbility Index of Charlson, oliguria, low serum albumin and cholesterol, and anemia were significantly associated with mortality (p<0,05). The lineal regression analysis found three factors associated to the mortality rate: Comorbility Index of Charlson, oliguria and low serum cholesterol. Mortality predictive model was carried out. CONCLUSION: Highest basal comorbility of patients, oliguria and malnutrition-inflammation dates are princess prognosis and mortality factors in ARF today A new approach is needed in ARF because this new type/class of population.

Doxazosina asociada a la combinación bloqueante del eje-renina-angiotensina y calcioantagonista en pacientes con insuficiencia renal crónica / Doxazosine associated to renin-angiotensin blockers and calcioantagonists in chronic renal failure patients

Objetivo. El estudio ZAFRA se diseñó para evaluar la seguridad de un nuevo bloqueantede los canales del calcio, lercanidipino, en la insuficiencia renal crónica, ysu posible sobre la función renal en los pacientes tratados con fármacos que bloqueanel eje renina-angiotensina. Los pacientes que no controlaron su PA con estos fármacosfueron tratados con doxazosina.Diseño y métodos. El estudio reclutó 203 pacientes con insuficiencia renal (creatinina> 1,4 mg/dl en varones o > 1,2 mg/dl en mujeres, o aclaramiento de creatinina< 80 ml/min). Todos los pacientes estaban siendo tratados con IECA o antagonistasde receptores y la medicación se mantuvo a lo largo del estudio, sin quepudieran recibir diuréticos a lo largo del seguimiento. Los pacientes eran evaluadosclínica y analíticamente 1, 3 y 6 meses después de iniciar tratamiento con lercanidipino.Aquellos pacientes que en la segunda visita no estaban controlados añadierondoxazosina GITS (4 mg en dosis única) al tratamiento.Resultados: 57 pacientes que iniciaron el estudio fueron tratados con doxazosina(edad media 64,8 ± 12,7 años, 47,4 varones y 52,6 mujeres). La PA se redujo significativamentedesde 164 ± 17/92 ± 9 hasta 135 ± 13/78 ± 8 mmHg (p < 0,001). Seprodujeron reducciones significativas de la PA en el 67,6% de los enfermos y el32,4% de los pacientes redujeron su PA hasta los límites recomendados (<130/85mmHg). Únicamente 2 (3,6) pacientes han presentado reacciones adversas. No sedetectó aumento de la incidencia de edema. La creatinina plasmática (1,9 ± 0,6mg/dl) no se había modificado al final del estudio (2,0 ± 0,8 mg/dl) ni el aclaramientode creatinina (38,7 ± 15,7 vs 38,5 ± 14,5). También se detectó un descensosignificativo del urato plasmático (7,0 ± 1,9 vs 6,7 ± 1,6 mg/dl, p < 0,05).Conclusiones: La doxazosina ha presentado un excelente perfil de seguridad enlos pacientes renales, destacando la ausencia de edemas a pesar de la insuficienciarenal de los pacientes

Objective. To evaluate the safety and effectiveness of the alfa-blocker doxazosinGITS in CRF patients.Design and methods. The study recruited 203 CRF patients (creatinine > 1,4mg/dl for males, creatinine > 1,2 mg/dl for females, or creatinine clearance < 80ml/min). All patients were receiving ACE inhibitores (63.4%) or angiotensin II antagonist(36.6%) therapy but they had higher blood pressure than recommended forCRF (130/85 mmHg). Patients were clinically evaluated 1, 3 and 6 moths after startingtreatment with lercanidipine (10 mg once daily). Patients with high blood pressurein spite of combined therapy with two drugs added doxazosin GITS 4-8 mgonce daily to treatment.Result: 57 patients rendered evaluable for the study (age 64.8 ± 12.7 years,47.4% males and 52.6 females). BP significantly decrease from 164 ± 17/92 ± 9mmHg to 135 ± 13/78 ± 8 mmHg. 67.6% patients showed a significant BP reductionand 32.4% gets optimal BP control (< 130/85 mmHg). Two patients (3.6%)showed untoward effects. No biochemical changes were detected.Conclusions: Doxazosin showed a good antihypertensive effect in CRF patientswhen used as third drug in resistant severe hypertension. It has a good tolerabilityprofile and showed a neutral profile on biochemical parameters

[Doxazosine associated to renin-angiotensin blockers and calcioantagonists in chronic renal failure patients]. / Doxazosina asociada a la combinación bloqueante del eje-renina-angiotensina y calcioantagonista en pacientes con insuficiencia renal crónica.

OBJECTIVE: To evaluate the safety and effectiveness of the alfa-blocker doxazosin GITS in CRF patients. DESIGN AND METHODS: The study recruited 203 CRF patients (creatinine > 1,4 mg/dl for males, creatinine > 1,2 mg/dl for females, or creatinine clearance < 80 ml/min). All patients were receiving ACE inhibitores (63.4%) or angiotensin II antagonist (36.6%) therapy but they had higher blood pressure than recommended for CRF (130/85 mmHg). Patients were clinically evaluated 1, 3 and 6 moths after starting treatment with lercanidipine (10 mg once daily). Patients with high blood pressure in spite of combined therapy with two drugs added doxazosin GITS 4-8 mg once daily to treatment. RESULT: 57 patients rendered evaluable for the study (age 64.8 +/- 12.7 years, 47.4% males and 52.6 females). BP significantly decrease from 164 +/- 17/92 +/- 9 mmHg to 135 +/- 13/78 +/- 8 mmHg. 67.6% patients showed a significant BP reduction and 32.4% gets optimal BP control (< 130/85 mmHg). Two patients (3.6%) showed untoward effects. No biochemical changes were detected. CONCLUSIONS: Doxazosin showed a good antihypertensive effect in CRF patients when used as third drug in resistant severe hypertension. It has a good tolerability profile and showed a neutral profile on biochemical parameters.

[Lercanidipine in diabetic patients with renal failure]. / Lercanidipino en pacientes diabéticos it con insuficiencia renal.

OBJECTIVE: To evaluate the safe use of a new calcium channel blocker, lercanidipine, in diabetic chronic renal failure (CRF) patients. DESIGN AND METHODS: The study recruited 42 diabetic CRF patients (creatinine > 1.4 mg/dl for males, creatinine > 1.2 mg/dl for females, or creatinine clearance < 70 ml/min). Mean age was 68.2 +/- 9.1 years. 53.8% were males and 46.2% females. Three patients were type 1 diabetics and 39 ones were type II. All patients were receiving ACE inhibitors (67.4%) or angiotensin II antagonist (32.6%) therapy but they had higher blood pressure than recommended for CRF patients (130/85 mmHg). No patients were under diuretic treatment. Patients were clinically evaluated 1, 3 and 6 months after starting treatment with lercanidipine. Samples for urine and blood examination were taken during the examination. When needed, a third drug was added to treatment, excluding diuretics. Creatinine clearance was measured using 24 h urine collection. RESULTS: BP significantly decrease from 163 +/- 18/90 +/- 8 mmHg to 134 +/- 12/77 +/- 9 mmHg. One half of patients showed significant reduction of blood pressure, 26.7% reached the target blood pressure (< 130/85 mmHg) and 20.0% gets optimal BP control (< 130/85 mmHg). No one patient showed untoward effects. No edema was detected nor adverse effects related to vasodilatation were found. Plasmatic creatinine did not change (1.9 +/- 0.5 baseline vs 1.8 +/- 0.5 mg/dl) and creatinine clearance increased at the end visit (40.1 +/- 14.5 baseline vs 45.4 +/- 18.2 ml/min) but the difference was not significant. Proteinuria was unchanged. CONCLUSIONS: Lercanidipine showed a good antihypertensive effect in diabetics CRF patients. It has a good tolerability profile and showed neutral effect on plasmatic lipids. Neither impairment of renal function nor increment in proteinuria were detected.

Decreased oxidative stress in prehepatic portal hypertensive rat livers following the induction of diabetes.

1. Oxidative stress (OS) is a biological entity indicated as being responsible for several pathologies, including diabetes. Diabetes can also be associated with human cirrhosis. Portal hypertension (PH), a major syndrome in cirrhosis, produces hyperdynamic splanchnic circulation and hyperaemia. The present study was designed to investigate the occurrence of OS in prehepatic PH rat livers following the induction of diabetes. 2. Five groups of rats were used: control, sham operated, chronic diabetes (induced with a single dose of streptozotocin at 60 mg/kg, i.p.), prehepatic PH and chronic diabetic plus prehepatic PH. The occurrence of OS was determined in liver homogenates by measuring hydroperoxide-initiated chemiluminescence and the activity of anti-oxidant enzymes (superoxide dismutase, catalase and glutathione peroxidase). 3. Prehepatic PH produced a significant increase in hydroperoxide-initiated chemiluminescence in the liver compared with control and sham-operated rats, whereas the liver in chronic diabetic rats showed no difference. However, chemiluminescence values decreased almost by 50% in the chronic diabetic plus prehepatic PH group. Concomitantly, the activities of the anti-oxidant enzymes in chronic diabetes, prehepatic PH and chronic diabetic plus prehepatic PH groups were decreased (P < 0.05 vs control and sham-operated groups). 4. Livers from the chronic diabetic group did not show any evidence of the occurrence of OS, whereas the prehepatic PH group showed the occurrence of OS. The association of PH and chronic diabetes resulted in a significant decrease in the occurrence of OS, which could be explained by an anti-oxidant response to an OS.