CHEMISTRY REAGENT KITS----->UREA NED KINETIC
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CLINICAL SIGNIFICANCE
Increased levels are associated with renal disease as well as dehydration, diabetic coma, hypoadrenal crisis, gastrointestinal hemorrhage and circulatory collapse. Decreased values are observed in some case of severe liver disease.

PRINCIPLE
In the presence of o-Phthalaldehyde and naphthylethylene diamine sulfate complex in an acid medium urea reacts with o-PA-NED complex to produce a pink colored compound, the rate of formation of this complex which can be monitored on initial rate reaction or fixed time kinetic mode at 505 nM ± 15 nM is in proportion to the urea concentration. The acidic reactive medium makes to avoid deproteinization of the serum.

COMPOSITION  
o-Phthalaldehyde 29 mMol/L

Contain acidic buffer and stabilizers

Naphthylethylene diamine sulfate   23 mMol/L

Contain acidic buffer and stabilizers

Urea 40.00 mGs/dL

Or

 
Urea Nitrogen 18.65 mGs/dL

REAGENTS SUPPLIED

 
1.Cat.No.LS398 4x 50 mL
o-PA R1 2x50mL
NED R2 2x50mL
UREA STANDARD

3 mL

Preparation of Reagent
All Reagents are ready to use.

STABILITY OF REAGENT
Store all reagent at 2-8°C. Reagents 1 and 2 are stable for 24Months. A pale yellow color will develop In reagents If contaminated with each other. Discard the reagent If a Brown-Yellow color Is developed.

SAMPLE
Sample can be serum or plasma which has no sign of hemolysis. Common anticoagulants have no Interference on this assay. Marked deterioration may not be found If samples stored In a refrigerator for 72 hrs. If urine sample to be tested It must be diluted 1 to 100 and multiply the result by 100.

 

METHOD FOR AUTOANALYSERS

   Temperature 37°C
   o-PA Reagent 1 250 µL
  Standard or Sample 50 µL
  NED Reagent 2 250 µL

Mix well and feed the analyzer in the sequence as given under.

  1. Zero with distilled water
  2. Feed standard assay and calibrate
  3. Feed test assays

NOTE:Programme the analyser using system parameters. A specific programme data sheet may be provided for each analyser upon request.

SYSTEM PARAMETERS


Program the analyzer as under

Mode =

Initial Rate or Fixed Time Kinetic

Blanking =

Distilled Water

Wavelength nM =

505(490-520)

Temperature =

37°C

Optical Path Length = 1 cM
Sample volume =

50 ni

Reagent 1 volume =

250 ni

Reagent 2 volume =

250 ni

Concentration of Std. =

40 mGs/dL

Delay time =

60 seconds

Interval =

60 seconds

Maximum ABS limit =

12.00

Unit = mGs/dL
Linearity = 120 mGs/dL

RESULTS
                          40
Factor = ------------------------
                 ΔA/min Standard

D A/min = O.D. 120 seconds - OD 60 seconds
40 = Concentration of STD. in mGs/dL

Samples having urea grater than 120 mGs/dL
must be reassayed with dilution in distilled water
and multiply the value with dilution facto i.e.2
for 1:1
BUN = Urea x 0.466
UREA mMol/L = mGs x 0.1665

EXPECTED VALUES
Serum or Plasma Urea up to 40mGs/dL
(18 mGs/dL BUN).
Urine Urea 20-35 Grams/24 hrs.

LIMITATIONS
This reagent system requires protein precipitation in case of sample having hyper-macroglobulinamea to avoid turbidity with working reagent resulting in false reporting and formation of potential mass of precipitated protein in teh flow cell. Therefore, such sample should be pre-identified and treated with suitable protein precipitants. o-PA-NED working reagent is susceptible to deterioration. This assay is not linear above 120 mGs/dL urea. Introducing contaminated pipettes or chemicals make false results. Low sensitivity of reaction is observed if reagents are contaminated with each other or working reagent prepared more than one hour earlier.

QUALITY CONTROL
To ensure adequate quality control, each run should be accompanied with a standard quality control serum. The accuracy of the test must be established against the values of control serum calibrated as per this methodology. It should be realized that the use of quality control material checks both instrument and reagent function together. Factors which might affect the performance of this assay include proper instrument functions, temperature control, cleanliness of glassware, accuracy of volumetric systems, timings and appropriate reagent storage & handling

WARNING
This reagent system is for in vitro Diagnostic use only. This reagent system is containing components that have not been established for safety if contacted on broken skin or eye or taken orally. In case of such incidents wash off with plenty of water.

BIBLIOGRAPHY
1.Levinson SS, Clin. Chem 24, 12, (1978)
2. Lequang CT.CIin. Chem 33, 192,(1987)