Haemoglobin – Part 1 – Hemoglobin (Hb)

Sample

1. The blood sample is taken in EDTA.
   1. EDTA salt of sodium or potassium is preferred sample.
2. The blood sample is stable 48 hours at 4°C and 24 hours at 23 °C.

Precaution

1. Avoid clotting (micro-clots may form) which will lower the Hb.
2. Falsely high results may occur due to prolonged venous stasis during venipuncture.
3. Increased turbidity, the presence of paraproteins, and nucleated RBCs gives high values.
4. Lipemic blood also gives high values.
5. In pregnancy, there is a false low Hb due to increased blood volume.
6. In the high altitude area, the Hb will be high.
7. Gentamicin and methyldopa may increase Hb value.
8. Antibiotics, chemotherapy, aspirin, and sulphonamide gives low values.

Purpose of the test (Indications)

1. This is done to diagnose anemia.
2. It tells the severity of anemia.
3. This will monitor the effectiveness of the treatment of anemia.
4. This is done to evaluate polycythemia.
5. This is a part of the complete blood examination.

Pathophysiology

1. Hb is a globular protein with a diameter of 6.4 nm in diameter.
2. Hb A has a molecular weight of 68,000  and one RBC contains 640 million Hb molecules RBC.
   1. Hb consists of four polypeptide chains α2 and β2  and heme groups. The 4 iron atoms are in the ferrous state (Fe++).
   2. Heme synthesis takes place in the mitochondria.
   3. Hb is 90% in the RBCs.

Age or stage of the fetus	Hb type	Structure
In early embryonic life	Gower 1	ζ2 ε2  (zeta and epsilon)
	Gower 2	α2 ε2  (alpha and epsilon)
At 6th weeks of gestation	Fetal Hb	α2 γ2  (alpha and gamma)
At 28 the week of gestation	Hb A	α2 β2  (alpha and beta)

Globin gene location on the chromosomes:

3. Hemoglobin synthesis is a complicated process. This takes place in the red blood cells.
   1. To understand, this is summarized in the diagram.
   2. Amino acids are assembled in the ribosomes and give rise to α2 β2 globulin.
   3. Transferrin gives Iron which will combine with the haem molecule.
   4. Heme molecule will have protoporphyrin by the chemical combination of glycine and succinyl CoA in the presence of Vitamin B6 and form heme.
      1. Heme molecule consists of protoporphyrin IX protein and iron (Fe++).
      2. Protoporphyrin + Fe++ = Heme.

4. Hb is the iron-containing pigment of the RBCs with the main function of carrying oxygen to various parts of the body.
   1. There is a central part of the Heme (iron-porphyrin)  group that is the site of Oxygen uptake and release.

5. Hb, serve as the main vehicle for the transportation of oxygen and carbon dioxide.
   1. It carries oxygen from the lung to tissues.
      1. CO2 from the tissue to the lung.

6. Hb consists of protein portion as Globin and iron part as heme.
7. Iron combines with oxygen and gives it a red color.
   1. The oxygen combining capacity is directly related to Hb concentration and not on the number of RBCs.

8. Hb acts as a buffer in the extracellular fluid (acid-base buffer system).
   1. In tissues, the oxygen concentration is lower and CO2 and H+ ions concentration are higher.
   2. When the pH is lower then oxygen dissociates from Hb.
   3. Now deoxygenated Hb will bind to H ions then raise the pH.
   4. CO2 diffuses into  RBCs and forms carbonic anhydrase, it will convert CO2 to  HCO3¯ and protons.
   5. Protons are bound to Hb  HCO3¯ ions leave the cell.
   6. For every HCO3¯ ion leaving the cell will lead to the entry of a chloride ion.
   7. This buffer system is dependant upon the lungs and kidney to eliminate CO2 and HCO3¯.
9. Abnormalities in the globin structure lead to hemoglobinopathies like Sickle cell anemia, and Hb C,
   1. In thalassemia, there is an abnormality in the globin chain synthesis.
10. Hb closely reflects the hematocrit and RBC count.
11. Hb at birth is usually lower in premature infants.
12. There is no significant change in Hb concentration after the age of 85 years.
13. The main function of hemoglobin is to transport oxygen to the tissues and remove carbon dioxide.

Sign and symptoms due to Hb level:

1. Low Hb:
   1. puts a strain on the cardiopulmonary system to maintain the oxygen level.
   2. There is a risk of angina, heart attack, congestive heart failure.
   3. stroke.
2. High Hb:
   1. the chances for intravascular settling.
   2. Stroke.
   3. Infarction in other organs.

Normal values

Source 1

Age	Hb g/dL
Fetal Hb
18 to 20 weeks	11.7  ± 0.78
21 to 22 weeks	12.28 ± 0.89
23 to 25 weeks	12.40 ± 0.77
26 to 30 weeks	13.35  ± 1.17
Cord blood	13.5 to 20.5
Infants
0.5 month	13.4 to 19.8
1 month	10.7 to 17.1
2 month	9.4 to 13.0
4 month	10.3 to 14.1
6 month	11.1 to 14.1
9 month	11.4 to 14.0
one year	11.3 to 14.1
Child
0.5 to 2 years	11.0 to 14.0
2  to 5 years	11.0 to 14.0
5 to 9 years	11.5 to 14.5
9 to 12 years	12.0 to 15.0
	Male	Female
12 to 14 years	12.0 to 16.0	11.5 to 15.0
15  to 17 years	11.7 to 16.6	11.7 to 15.3
Adult
18 to 44 years	13.2 to 17.2	11.7 to15.5
45 to 64 years	13.1 to 17.2	11.7 to 16.0
65 to 74 years	12.6 to 17.4	11.7 to 16.1

• To convert into SI units x 10 = g/L

Another source

1. Women = 12 to 16 g/dl
2. Men = 14 to 17.4 g/dl
   1. Pregnant women = > 11 g/dl.

Another source

Normal values in Infant and Children

0 to 2 weeks	14 to 24.5 g/dl
2 to 8 weeks	12.5 to 20.5 g/dl
2 to 6 months	10.7 to 17.3 g/dl
1 to 6 years	9.5 to 14.1 g/dl
6 to 16 years	10.3 to 14.9 g/dl

Normal value in Cord blood

• 9 month = 11.4 to 14 g/dl

• The normal values should be decided according to the population of various countries.

Normal values of a different fraction of Hb in the adult:

• Hemoglobin A 2  =  1.5 to 3.5 % of the total Hb.
• Hemoglobin F  = < 1% of total Hb.
• Hemoglobin in plasma  = 0.5 to 5.0 mg/dL.
• Methemoglobin  = < 1% of total Hb.

Critical values

1. Blood transfusion is not recommended as long as the Hb is above 8 g/dl. and Hct is above 24 %.
2. Hb < 5 g/dl is a very critical level and needs a blood transfusion.
3. In older patients, blood transfusion is recommended when the Hb level is below 10 g/dl.

Increased Hb is seen in:

1. Polycythemia.
   1. Polycythemia vera.
   2. congestive heart failure.
   3. Chronic obstructive pulmonary disease (COP).
2. After vigorous exercise.
3. Hemoconcentration like dehydration, burns, severe vomiting.
4. Intestinal obstruction.
5. Severe dehydration like diarrhea, and burns.

Decreased Hb is seen in:

1. Anemia.
2. Drugs that cause aplastic anemia.
3. Drugs that cause hemolysis ( G6PD deficiency ).
4. Immune hemolytic anemia.
5. Iron deficiency anemia.
6. Thalassemia.
7. Pernicious anemia.
8. Hemoglobinopathies.
9. Liver diseases, and Cirrhosis.
10. Hypothyroidism.
11. The hemorrhage acute or chronic like bleeding hemorrhoids, or ulcers.
12. Malignancies:
    1. Hodgkin’s disease.
    2. leukemia.
    3. Lymphomas.
    4. Carcinomatosis.
    5. Multiple myelomas.
13. Autoimmune diseases.
    1. SLE.
    2. sarcoidosis.
    3. Rheumatoid arthritis.
14. Dietary deficiency

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