Hemoglobin

Electrophoresis
Hemoglobin Structure
The subunit composition & percent of the
principal normal hemoglobins are shown in the
following table:

Fraction of total Chain Form

hemoglobin composition
22 95 – 98% HbA
22 < 2% HbF
22 2 – 5% HbA2
 
Hemoglobinopathies
Hemoglobinopathies have traditionally been defined as a
family of disorders caused by production of a :
(i) Structurally abnormal hemoglobin molecule
(ii) Synthesis of insufficient quantities of normal
hemoglobin, or,
(iii) Rarely both.
Sickle-cell anemia (HbS), hemoglobin C disease (HbC),
Hb M, and the thalassemia syndromes are representative
hemoglobinopathies that can have severe clinical
consequences.
The first three conditions (HbS, HbC &
HbM)result from production of hemoglobin
with a mutation in the α and/or β genes leading
to altered amino acid sequence, whereas the
thalassemias are caused by decreased
production of normal hemoglobin.
 A. Sickle cell disease (HbS disease)
Sickle cell disease (also called sickle cell anemia) is a
genetic disorder of the red blood cells caused by a single
nucleotide alteration (a point mutation) in the β-globin
gene.
A molecule of HbS contains two normal α-globin chains
and two mutant β-globin ( βs) chains in which the polar
glutamate at position six has been replaced with a
nonpolar valine.
 Homozygotes have HbS but no HbA.
Heterozygotes have both hemoglobins in their red
cells, so they do not suffer from the disease and are
only carriers (sickle cell trait).
 1 2 3 4 5
Lane 1: Sickle cell trait (AS) (Heterozygous)
Lane 2 & 3:Sickle cell disease (SS) (Homozygous)
Lane 4: Normal (AA) (Normal Adult)
Lane 5: Control
 1 2 3 4

A
F

Lane 1: Control
Lane 2 &4: AS (Sickle cell trait)
Lane 3: SS (Sickle cell disease)
 B.Thalassemias

The thalassemias are hereditary diseases in

which there is defective production of either
α-chain (α-thalassemia) or the β- chain (β-
thalassemia).
A thalassemia can be caused by a variety of
mutations, including entire gene deletions, or
substitutions or deletions of one to many
nucleotides in the DNA.
1- α-Thalassemias:
These are defects in which the synthesis of α -
globin chains is decreased or absent.
2- β-Thalassemias:
In these disorders, synthesis of β-globin chains is
decreased or absent, whereas α-globin chain
.synthesis is normal
 β-Thalassemias
There are only two copies of the β-globin gene in
each cell (one on each chromosome 11).
Therefore, individuals with β-globin gene defects
have either β-thalassemia trait (β-thalassemia
minor) if they have only one defective β-globin
gene, or β-thalassemia major if both genes are
defective.
 HbA

HbA2

a) Beta thalssemia major showing strong band at HbF and

HbA2 position
b)Normal control showing strong band at HbA position
c )Beta thalassemia trait showing strong band at HbA and
visible band at HbA2 position which is stronger than that of
normal control
Hemoglobin Electrophoresis Patterns in Beta Thalassemia
Lanes 1 and 2:normal patient specimen
Hb A is over 98% with a small amount of Hb A2 visible
Lanes 3 and 4: Beta thalassemia minor
Hb A is decreased to 94%, Hb A2 is increased at 5%, and Hb F
is 1%
Lanes 5 and 6: Beta thalassemia major
No Hb A or A2 is present, Hb F is 100%