Yazhid Blog


Senin, 20 Maret 2017


Photometric colorimetric test for determination of total protein, biuret method

Cupric ions react with protein in alkaline solution to form a purple complex. The absorbance of this complex is proportional to the protein concentration in the sample.

Reagent preparation and stability
R1 (colour reagent) and 2 (standard) are ready for use. They are stable even after opening up to the expiry date when stored at 2...25oc. Contamination  after opening must be avoided.

Serum. Heparinised or EDTA plasma

Stability in serum
Up to 1 mounth at 2...80c, up to 1 week at 15...250c.

Wavelength              : Hg 546 nm
Optical path              : 1 cm
Temperature           : 20...250c
Measurement         : againts reagent balnk.
                        Only one reagent blank per series is required.

Pipeting scheme
Pipette into cuvettes
Reagent blank
Sample or standard
Sample / standard
Mix, incubate for 10 min. At 20...250c. Measure the absorbance of the sample and the standard againts the reagent blank within 30 min.
The test is linear up to a protein concentration 12 g/dl or 120 g/l. Dilute samples with a higher concentration 1+1 with physiological saline (0,9 %). Multiply the result by 2.

Normal range in serum or plasma
Normal born babies                                              4,6-7,0 g/dl or 46-70g/l
Children from 3 years and adults     6,6-8,7 g/dl or 66-87 g/l

Quality control
For internal quality purpose, all cemmercially available control sera with total protein values determined by the biuret method may applied.

The test may be performed manually or on clinical analyzers. For applications please contact us at the address given below.

1.       The sample blank for clear, colorless sera is equivalent to 0,2 g/dl and can therefore be ignored. A sample blank must be determined for haemolytic and lipemic sera by pipetting 20µl sample to 1000µl  physiological saline and measuring againts dist, water. The absorbance of the sample blank has to be subtracted from the absorbance of the sample.
2.       The colour reagent contains sodium hidroxide which is irritant (R 36/38). In case of contact with skin and mucous membranes wash with copious amounts af water.
3.       The standard contains sodium azide as preservative (0,095 %) do not swallow. Avoid contact with skin and mucous membranes.
4.       A slight sediment may develop as the reagent ages. Do not include this sediment in the reaction mixture

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