3.1. Qualitative detection and quantitative determination of protein in the urine.

Normally, human urine contains a minimal amount of protein (an adult from 20 to 60 mg, children - from 3 to 47 mg per 24 hours), which presence cannot be proved by reactions generally used in laboratory. In a number of diseases amount of protein in the urine increases, and it begins to be determined by conventional methods. Color reactions are not suitable for the determining of the protein in the urine because of urine has its own color. In addition, it may contain substances that prevent the appearance of the characteristic color.

3.1.1. The test by boiling in weak acid environment.

In the test tube add 1 ml of urine and test its reaction to litmus. If the urine is alkaline, it is very weak acidified by adding 1% acetic acid. Urine is boiled. In the presence of protein turbidity or sediment is formed. During boiling the urine phosphates and carbonates of calcium and magnesium precipitate can be precipitate. In this case, the precipitate is dissolved by adding 1-2 drops of 10% acetic acid solution and it is again boiled.

3.1.2. The test by boiling in an acid medium in the presence of saturated sodium chloride solution.

To 1 ml of urine add 5 drops of saturated sodium chloride solution, 1-2 drops of 10% acetic acid solution and boil it. In the presence of protein a white flocculent precipitate or turbidity is formed.

3.1.3. Geller’s test.

In the test tube 1 ml of concentrated nitric acid is placed and gently from a pipette, lowering the wall of the tube, urine is added. There is a white amorphous precipitate, or turbidity in the form of a ring in the presence of protein at the boundary of two liquids. The test with concentrated nitric acid is sensitive - it opens up to 0.033 g/l of protein.

3.1.4. The test with sulfosalicylic acid.

Check the reaction of urine. Alkaline urine is acidified with 2-3 drops of 5-10% acetic acid solution. In the test tube pour 3 ml of urine and add 6 drops of 20% solution of sulfosalicylic acid. Urine that contains protein grows turbid. The results are read on a dark background. Test with sulfosalicylic acid is one of the most sensitive; it opens the protein in its content in the urine up to 0.015 g/liter. However, the test gives false positive results with certain medications (penicillin derivatives).

3.1.5. Quantitative determination of protein in the urine by the method of dilution (Brandberg - Roberts - Stolnikov method).

Experimentally established that if there is the interaction of a nitric acid layer and a urine layer, and the formation of protein ring will occur between 2 and 3 minutes, it shows that the urine contains 0.033 g/l of protein.

In the test tube 1 ml of Larionov’s reagent or nitric acid is placed and the same amount of urine is carefully poured on the wall of the tube. In considering the emergence of thin rings of protein between the 2^nd and 3^rd minutes of the boundary between two liquids on the dark background indicates the presence in urine 0.033 g/l of protein. If the ring appears before the two minutes after layering, the urine should be diluted with water and repeate the test. At the same time choose a special dilution of urine, during a layering of which the ring is formed between the 2^nd and 3^rd minutes. If the ring has a strand-like form, the urine should be diluted 2-fold, with the broad shape of a ring - 4-fold and with a compact form 8-fold. The amount of protein is calculated by multiplying 0.033 g/l on the dilution.

The calculation of the amount of protein, depending on the dilution of urine is presented in Table 7.

Table 7