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Test principle
The test paper is impregnated with tetramethylbenzidine and an organic hydroperoxide. The peroxidative effect of the haemoglobin and myoglobin oxidises the chromogen to form a blue-green dye, which causes the yellow test paper to turn green.
Intact erythrocytes lyse on the test pad. The haemoglobin released by the cells causes a change in colour, visible green dots appear in the surroundings of the erythrocytes. The haemoglobin dissolved in urine (lysed erythrocytes) gives a homogeneous green colour.
Sensitivity
The test pad detects approx. 0.015 - 0.03mg free haemoglobin/dl urine.The practical sensitivity to erythrocytes is 5 to 10 erythrocytes/µl.
Specifity
The test specifically reacts to haemoglobin and myoglobin.
Sources of errors
  • Nonspecific oxygen receptors such as ascorbic acid in high concentrations (e.g. following medication with vitamin C, uric acid, glutathione and gentisic acid) can diminish the sensitivity of the test.
  • False-positive results can be caused by hypochlorite, residual highly oxidating cleaning agents or disinfectants in the urine container.
  • High concentrations of nitrite (above 10 mg/dl, 2.2 mmol/l) reduce the speed of the reaction.
  • Formalin should not be used for preserving urine as too low or falsenegative results can result.
  • High protein concentrations (proteinuria, > 500 mg/dl, > 5 g/l) can reduce the colour intensity of the reaction.
  • False-positive results can occur due to microbial peridoxase activities in urogenital tract infections.
Clinical significance
Haematuria, the excretion of intact erythrocytes in urine, is always a serious symptom and requires careful clarification of the cause. In haemoglobinuria, free haemoglobin (lysed erythrocytes) occurs in urine. Myoglobinuria is usually caused by injured muscles or muscle necrosis (myonecrosis).Macrohaematuria can be detected with the naked eye due to the red coloration of the urine (blood contents > 0.5 ml blood/l, > 2500 erythrocytes/µl). Certain medicines (such as pyrazolones, phenazopyridines) or dyes (such as after eating beetroot) can also cause a red coloured urine. Haematuria can occur in many diseases. These mainly concern: 
  • diseases of the kidneys,
  • diseases of the urogenital tract
  • haemorrhagic diatheses.
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