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Urea ct fs

Manufactured by DiaSys
Sourced in Germany

DiaSys Urea CT FS is a diagnostic reagent kit used for the quantitative determination of urea concentration in human serum, plasma, or urine samples. The test is based on the enzymatic hydrolysis of urea by urease, and the subsequent reaction of the released ammonia with 2-oxoglutarate and NADH in the presence of glutamate dehydrogenase. The decrease in NADH absorbance is measured photometrically and is proportional to the urea concentration in the sample.

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6 protocols using urea ct fs

1

Measurement of Urine and Plasma Biomarkers

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Urine albumin was measured with a mouse albumin ELISA (Bethyl Laboratories Inc. Montgomery, TX). Sodium and potassium excretion were determined by flame photometry. Plasma urea was measured by DiaSys Urea CT FS (DiaSys Diagnostic Systems, Holzheim, Germany).
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2

Plasma Biomarkers of Organ Dysfunction

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Blood extraction was performed by cardiac puncture into lithium heparin-containing tubes. The blood was immediately centrifuged at 4,000 rpm for 10 min at 4 °C to isolate the plasma. To measure markers of liver and kidney failure, we quantified plasma levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), total bilirubin (TBIL) for liver function, and creatinine (CRE) and blood urea nitrogen (BUN) for kidney function. Markers were quantified using the Piccolo Xpress Chemistry Analyzer (General Chemistry 13, MetLyte Plus CRP and Basic Metabolic Panel Plus panels, Abaxis, USA) according to manufacturers' instructions. Plasma urea was determined by DiaSys Urea CT FS (DiaSys Diagnostic Systems, Holzheim, Germany), according to the manufacturers’ instructions. Plasma levels of kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin (NGAL) and β2-microglobulin (β2M) markers were quantified by corresponding rat ELISA Kit (Abcam).
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3

Assessing Renal Function in Transplant Rats

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We performed tail-cuff SBP registration and collected 24-h urine samples for determination of protein excretion (Bradford Protein Assay, Bio-Rad Laboratories, Hercules, CA, USA) with the rats in individual metabolic cages while fasting, as described (Bongartz et al., 2010 (link)), at weeks 3 and 5 after transplantation. Blood samples were collected from the tail vein at the same time-points for determination of plasma urea (DiaSys Urea CT FS, DiaSys Diagnostic Systems, Holzheim, Germany).
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4

Surgical Induction of Chronic Kidney Disease

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CKD was induced by surgical 5/6th nephrectomy (SNX) as published,36 (link) with minor adaptations. In short, in a single procedure, surgical removal of the whole left kidney and polectomy (2/3rd) of the right kidney was performed using retroperitoneal incisions. In Sham animals, both kidneys were externalized and the renal capsule was removed. Postoperative buprenorphine (0.03 mg/kg Temgesic) was given subcutaneously for a period of at least 36 h. Rats were weighed 3 times a week, 24-h urines, and blood samples were collected and the SBP was measured (Tail cuff sphygmomanometer–LE 5002 LETICA®) biweekly. To collect 24-h urines, rats were placed in metabolic cages without food for 24 h, but with free access to water with 2% glucose. Urine was collected in antibiotic/antimycotic solution (A5955; Sigma, St Louis, US) and stored at −20 °C. Blood samples were collected from the tail vein into EDTA Microtainers (BD #365974). Urinary protein levels were measured by Bradford Assay (Bio Rad). Plasma urea was determined by DiaSys Urea CT FS (DiaSys Diagnostic Systems). Plasma creatinine levels were determined by DiaSys Creatinine PAP FS kit (DiaSys Diagnostic Systems,). eGFR was calculated with a plasma creatinine- and urea-based equation specific for rats.37 (link) We determined CKD as established once a threshold of proteinuria exceeding 50 mg/24 h was reached.
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5

Metabolic and Cardiovascular Profiling in Rats

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Rats were weighed weekly, and at regular intervals 24-hour urine and blood samples were collected and the systolic blood pressure was measured by tail cuff sphygmomanometry at weeks 5, 9 and 11. To collect 24-hour urine, rats were placed in metabolism cages without food for 24 hours, but with free access to water with 2% glucose. For the systemic in vivo pravastatin treatment studies, pravastatin was also supplemented to the drinking water during urine collection. Urine was collected in antibiotic/antimycotic solution (A5955; Sigma) and stored at −80°C. Blood samples were collected from the tail vein. Urinary protein levels were measured with Coomassie blue. Sodium and potassium levels were determined by flame photometry. NO metabolites were measured (Cayman Chemical, Ann Arbor, MI, USA). Plasma urea and plasma and urinary creatinine levels were determined by DiaSys Urea CT FS (DiaSys Diagnostic Systems, Holzheim, Germany). Creatinine clearance was calculated by dividing urine creatinine excretion (μmol/minute/100 g body weight) by plasma creatinine (μmol/μl). Cholesterol and triglycerides were determined by DiaSys Cholesterol FS and DiaSys Triglycerides FS (DiaSys Diagnostic Systems).
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6

Quantification of Kidney Injury Markers

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After saline, HDL and oxHDL administration, plasma levels of creatinine (CRE), blood urea nitrogen (BUN) were quantified using a Piccolo Xpress Chemistry Analyzer (Abaxis, CA, USA), according to the manufacturers' instructions. Plasma urea was determined by DiaSys Urea CT FS (DiaSys Diagnostic Systems, Holzheim, Germany), according to the manufacturers' instructions. Plasma levels of KIM-1, NGAL and β2M markers were quantified by corresponding rat ELISA Kit (Abcam).
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