Urine-Hemoglobin in the urine (Hemoglobinuria)

Sample

1. It can be done with fresh, random urine.
2. A morning sample is preferred.
3. Examine immediately.
4. If examined later on say for one hour then refrigerate the urine in the fridge to avoid microbial growth.
5. Mix urine well before testing the urine for Hb.

Indications

1. To find the cause of hemoglobinuria due to various conditions:
   1. Hemolytic anemia.
   2. other conditions are given below.

Precautions

1. Examine immediately.
2. Can refrigerate if delayed.
3. Mix well before testing.

Pathophysiology

1. Definition of hemoglobinuria:
2. This is the presence of free hemoglobin in the urine, called hemoglobinuria.
   1. When the level of free hemoglobin in the blood exceeds the renal excretion threshold, then it results in hemoglobinuria.
   2. This lysis of the RBCs takes place in dilute, alkaline urine.
   3. The lysis of the RBCs in the urine usually shows a mixture of hemoglobinuria and hematuria.
   4. Urine in hemoglobinuria and myoglobinuria is dark red or brown.
      

      Causes of hemoglobinuria

3. Mechanism of hemoglobinuria:
   1. Erythrocytes lysis occurs at specific gravity <1.010.
   2. This may occur as a result of lysis of RBCs in the urinary tract.
   3. Intravascular hemolysis leads to hemoglobinuria. In such cases, no RBCs are seen.
   4. This will take place when the reticuloendothelial system can not handle or metabolize the hemoglobin by the hemolysis of RBCs (intravascular).
      1. Under normal conditions, there is a complex of haptoglobin-hemoglobin which will be not filtered by the glomeruli.
      2. When the hemoglobin exceeds the amount of haptoglobin in:
         1. Hemolytic anemias.
         2. Severe burns.
         3. Blood transfusion reactions.
         4. Strenuous exercise.
         5. Infections
      3. The free hemoglobin can pass through the glomeruli.
      4. The excess is absorbed in the proximal convoluted tubules.
      5. The free hemoglobin can filter out and appears in the urine as hemoglobinuria.
         

         Hemoglobinuria mechanism

   5. This may also take place by the hemolysis in the urinary tract system.
4. When RBCs are present in the urine the condition is called Hematuria.
   1. The presence of myoglobin is called myoglobinuria. This is due to myoglobin, a muscle protein.
      Ammonium sulfate test to differentiate hemoglobinuria and myoglobinuria:
      

      Urine ammonium sulfate test

5. Detection of hemoglobinuria:
   1. The small amount of hemoglobin in urine does not give visible color which can be detected by the dipstick method.
   2. O-toluidine gives a blue color with hemoglobin.
      1. This also detects myoglobin.
      2. The false-positive test is seen in:
         1. If the container contains hypochlorite, an oxidizing agent.
      3. The false-negative test is seen in:
         1. Vitamin C (ascorbic acid).
         2. Formaldehyde.
         3. Sensitivity is low when there are high specific gravity and a high concentrate of nitrite.
   3. Benzidine test is not used because this is a carcinogenic agent.
      

      Characteristics	Orthotolidine method	Benzidine method
      Substances detected	Hemoglobin Myoglobin	It is carcinogenic so not used now.
      False-negative result	Vitamin C (Ascorbic acid) Formaldehyde High specific gravity Increased nitrite concentration
      False-positive result	Hypochlorite in the container (detergents)

6. In case of positive urine for occult blood but no RBCs are seen under the microscope, then suspect myoglobinuria.
   1. This occurs as a result of muscle injury, muscle disease disorder, and some kind of poisoning.
7. This test detects:
   1. RBCs.
   2. Hemoglobin.
   3. Myoglobin.
8. Blood in the urine indicates damage to kidneys or the urinary tract.

Normal value

Source 1

• Normally Hemoglobin is negative in the urine.
• <0.03 mg free Hb/dL.

Causes of Hemoglobinuria:

1. It is positive for all causes of hematuria e.g
   1. renal calculi.
   2. tumors.
   3. exposure to toxic drugs.
2. Intravascular hemolysis.
   1. Pregnancy and in puerpera.
   2. Transfusion reaction.
   3. Kidney infarction.
   4. Extensive burns.
   5. Paroxysmal nocturnal hemoglobinuria.
   6. Drugs and poison e.g. sulfonamides, quinine, phenylhydrazine, poisonous snake, and fava beans.
   7. Infections e.g. malaria, blackwater fever, gas gangrene, yellow fever, and anthrax.
   8. Direct trauma to RBC like exercise, heart valve prosthesis.
   9. Microangiopathic hemolytic anemia.
   10. Disseminated intravascular coagulopathy.
   11. Hemolysis in the donor blood due to improper storage.Urine differentiation between the presence of RBCs, Hemoglobin, and Myoglobin:
       

       Characteristics tests	Hematuria	Hemoglobinuria	Myoglobinuria
       Reagent strip for blood	Positive	Positive	Positive
       Urine color	Cloudy red	Red, clear	Clear, red-brown
       Plasma color	Normal	Pink to red	Normal
       Presence of RBCs	Present	Absent (± few)	Absent (± few)
       LDH	Normal	Raised	Raised
       LD4 and LD5	Normal	Normal	Raised
       LD1 and LD2	Normal	Raised	Normal
       Total CK	Normal	Slight increased (10 times the normal)	Markedly increased (40 times the normal)

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