Urea test. Information about test and diagnosis of diseases

Dr. Yousef Al Adbai. 

                          UREA

1. Purpose: Quantitative estimation of urea in human serum by Urease-GLDH/UV kinetic method. Determination of serum urea nitrogen is an important index of kidney function. Impaired renal function or increased tissue protein breakdown is associated with increased urea nitrogen levels, whereas liver damage or pregnancy is associated with decreased levels.

2. Principle: Urea is hydrolyzed by urease to form ammonium carbonate. In the second reaction 2-oxoglutarate reacts with ammonium ion in the presence of glutamate dehydrogenase (GLDH) and the coenzyme NADH to produce L-glutamate. In this reaction two moles of NADH are oxidized to NAD^+ for each mole of urea hydrolyzed. The rate of decrease in the NADH concentration is directly proportional to the urea concentration in the specimen. It is determined by measuring the absorbance at 340 nm.

   

3. Performance specifications:

   · 3.1. Linearity: Up to 240 mg/dL of serum

   · 3.2. Measurement range: 2–240 mg/dL

   · 3.3. Sensitivity: Lower limit of detection is 2 mg/dL

4. Primary sample:

   · 4.1. Use plasma

   · 4.2. Collect 2 mL of venous blood from a peripheral vein in a heparin vasculature tube

   · 4.3. Do not use hemolyzed/contaminated plasma for testing

5. Type of container and additive

   · Use heparin/plain vasculature tubes for collecting samples; do not use hemolyzed/contaminated plasma for testing

6. Instrument: Semi-automatizer

7. Reagents/Consumables: The reconstituted reagent contains the following:

   · 7.1. TRIS pH 7.8, 2-Oxoglutarate, ADP, Urease, GLDH

   · 7.2. NADH

   · 7.3. Urea (50 mg/dL)

8. Procedure:

   · 8.1. Switch on the machine and press “FLUSH” button by keeping the tubing in a container with 2% detergent for 2 minutes followed by distilled water for 2 minutes.

· 8.2. Press “PROC”, different test procedures will be displayed.

· 8.3. Select the test to be processed by entering its number and then press “ENTER key.”

· 8.4. Now the assay parameters of the specific test procedure will be displayed. Note down the volume of the reagent and the sample to be used.

· 8.5. Run the standard with each batch of patient sample.

· 8.6. Then feed the test samples and record the values.

  · Assay: 2-point Kinetic

  · Sample volume: 10 μL

  · Wavelength: 340 nm

  · Reagent volume: 400 μL

  · Start reagent 100 μL

  · Temperature: 37 °C

9. Biological reference range: 15–38.5 mg/dL

10. Alert/Critical values: Above 80.0 mg/dL

11. Laboratory interpretation: Increase suggests impaired renal function, acute nephritis, chronic glomerulonephritis.

12. Potential sources of variability

   · 12.1. Use of only clear, unhemolyzed plasma separated from the erythrocytes as soon as possible. Lysed plasma specimens may give falsely elevated values

   · 12.2. On storage, the working reagent may develop a pink color which makes the use of reagent blanks necessary with every run.

   · 12.3. This method is recommended to perform only on mechanized equipment. It is difficult to include all samples and reagent blank exactly for the same intervals.

   · 12.4. The scheme may use for adaptation purpose for instruments with no specific adaptation sheet.

BIBLIOGRAPHY

1. Kassiter JP. New Eng J Med 1971;285:385.

2. Mackay EM, Mackay LL. Clin.Invest 1927;4:295.

3. Talke HN, Schubert, GE Kin. Wschr 1965;42:174.