What Is Hemoglobin Electrophoresis?

Hemoglobin electrophoresis is a blood test commonly used to diagnose and characterize disorders of hemoglobin, called hemoglobinopathies.

Hemoglobin is a complex protein, contained within the red blood cells, whose job is to carry and deliver oxygen throughout the body. Hemoglobin picks up oxygen from your lungs, transports the oxygen through your bloodstream, and releases it at the right time so it can be used by all the body's tissues.

There are several inherited hemoglobinopathies that can affect the ability of hemoglobin to perform its job normally.

Purpose of the Test

The hemoglobin electrophoresis test is designed to detect genetic abnormalities in the structure of a person's hemoglobin. Hemoglobin electrophoresis is typically done when a person has signs or symptoms of hemolytic anemia, a family history of a hemoglobinopathy, an abnormal complete blood count (CBC) test, or a positive neonatal screening test.

Currently, in the U.S., the American College of Obstetricians and Gynecologists recommends screening for hemoglobinopathy for all pregnant women by performing CBC testing, followed by a hemoglobin electrophoresis if the CBC is abnormal; or if the woman is at higher risk for a hemoglobinopathy based on ethnicity.

Women of African, Mediterranean, Southeast Asian, West Indian, or Middle Eastern descent are at higher risk for hemoglobinopathies. The male partners of women who are found to have hemoglobinopathies should also have screening if they are considering having a child.

The hemoglobin electrophoresis test is a blood test that can detect normal and abnormal hemoglobins, and begin to characterize the type of hemoglobinopathy if any exists. However, hemoglobin electrophoresis is only one of several tests that can detect and characterize abnormal hemoglobins.

Often, if an abnormal result is obtained with the electrophoresis test, sophisticated testing can be done to more precisely characterize hemoglobinopathies.

Hemoglobin electrophoresis is done by placing a small amount of blood on special paper or a special gel and exposing it to an electrical current. Different globins have different electrical charges and can be differentiated from one another based on their behaviors when exposed to an electrical current.

Different types of globin will move across the paper (or gel) at different speeds, and will thus separate themselves out into characteristic bands. By examining the bands that form during the application of the current, the types of hemoglobin present in the blood sample can be differentiated.

Risks and Contraindications

The hemoglobin electrophoresis test is a blood test. Consequently, there is almost no risk, aside from the small risk of bruising, bleeding, or infection that is present with any test requiring the drawing of blood.

In general, it is best not to have the hemoglobin electrophoresis test within 12 weeks of a blood transfusion, because it is possible for results to be confounded by hemoglobins from the transfused red blood cells.

Before the Test

The type of hemoglobin in your blood is not related to the time of day or what you have eaten or drunk lately, so there are no special instructions or restrictions you will need to follow before having a hemoglobin electrophoresis. The sample can be taken in any facility that performs standard blood drawing, at any time of day. Typically it is done in a healthcare provider's office, a lab, or a hospital. As with any blood test, you should wear comfortable clothing with loose sleeves that can be pulled up easily to expose your arm.

The hemoglobin electrophoresis test is usually covered by health insurance, as long as the healthcare provider provides the insurer with a reasonable explanation of why the test is indicated. However, it is always best to check with your insurance carrier before having the test, just to make sure. You should bring your insurance card with you when you have the blood test done.

During the Test

The hemoglobin electrophoresis test is done with a standard blood draw. A tourniquet will be placed on your arm and a technician will feel for a suitable vein. Your skin will be cleaned with an alcohol wipe, and a needle will be inserted into the vein and the blood sample taken. After blood is drawn, a small bandage or gauze patch will be applied. You will then be allowed to go home. 

After the Test

Complications from a blood draw are extremely unusual. Keep an eye out for any further bleeding, bruising, or inflammation or infection. If bleeding should occur, place more pressure on the puncture site for 5 to 10 minutes, and if the problem persists, call your healthcare provider. You should also call your healthcare provider if you see signs of inflammation or infection (redness, tenderness, excessive pain, or swelling).

Interpreting Results

You should expect to hear the results of your hemoglobin electrophoresis test within a few days to a week. If your test is normal, it may be that that's all you will hear.

However, you may get a more detailed report—or you may ask for a detailed report—even if the test is normal. 

Normal Hemoglobin Values

In adults, normal values for hemoglobin molecules are given as percentages, as follows:

• Hemoglobin A: 95%–98%
• Hemoglobin A2: 2%–3%
• Hemoglobin F: 0.8%–2%
• Hemoglobins S, C, D, E, and others: 0%

In children, higher levels of hemoglobin F are typical, with correspondingly lower levels of hemoglobin A and A2:

• Hemoglobin F in newborns: 50%–80%
• Hemoglobin F up to 6 months: 8%
• Hemoglobin F over 6 months: 1%–2%

Abnormal Results

If you have any amount of an abnormal hemoglobin on your hemoglobin electrophoresis, you will need further evaluation. 

Your healthcare provider will have to take into account many additional factors when interpreting the significance of the abnormal hemoglobin, including your family history, the results of your CBC (including particularly the hemoglobin, hematocrit, and the mean corpuscular volume), the appearance of your red blood cells under the microscope, and the results of your serum iron studies.

In addition, your healthcare provider may employ more sophisticated techniques to fully characterize and quantify the abnormal hemoglobin in your blood samples. Such testing may include high-pressure liquid chromatography, capillary zone electrophoresis, isoelectric focusing, or targeted genetic testing.

Understanding Hemoglobin and Hemoglobinopathies

Each hemoglobin molecule is a complex structure consisting of four protein subunits called globins, each of which is bound to a non-protein, iron-containing structure called a heme group. The four globin units in a hemoglobin molecule consist of two alpha-like and two beta-like chains. 

Each globin unit carries a heme group comprised of a porphyrin ring and an iron ion. It is the job of the heme group to bind and carry oxygen, and to release it to the peripheral tissues at the right time. Each hemoglobin molecule can bind four oxygen molecules.

The ability of hemoglobin to bind to oxygen—which is referred to as the oxygen affinity of hemoglobin—is largely determined by the globin subunits of the hemoglobin complex. Based on local environmental factors (especially the acidity of the blood and the local concentration of oxygen), the globin subunits change their shapes, as needed, to alter the affinity of their respective heme groups for oxygen. This calibrated oxygen affinity of hemoglobin allows oxygen molecules to be picked up, and then released, at just the right times.

As blood circulates through the lungs, oxygen diffuses into the red blood cells. In the local environment of the lungs, the oxygen is avidly taken up and bound by hemoglobin molecules. The oxygen-bearing hemoglobin is then carried out to the tissues. As the hemoglobin is exposed to an increasingly acidic environment in the peripheral tissues (caused by carbon dioxide waste produced by cell metabolism), it loses some of its affinity for oxygen. The oxygen is thus released to the tissues. 

The newly deoxygenated hemoglobin in the peripheral tissues picks up some of the excess carbon dioxide it finds there and carries it back to the lungs. (Most of the waste carbon dioxide, however, reaches the lungs after it is dissolved in the blood.)

Hemoglobin accounts for the color of blood. Hemoglobin in the arteries, carrying plenty of oxygen, is bright red in color (which is how red blood cells got their name). Hemoglobin in the veins, having delivered its oxygen to the tissues, becomes more blueish in color.

Types of Normal Hemoglobin

There are several types of hemoglobin, which are characterized by the specific types of globins they contain. Normal adult hemoglobin consists of two alpha and two beta globins. Other types of hemoglobins contain similar globins, often referred to as alpha-like and beta-like globins.

Three distinct kinds of hemoglobin are normally carried by red blood cells during different stages of human development. These three normal hemoglobins are optimized for their environment.

In very early gestation, when the human embryo receives its oxygen from the yolk sac, embryonic hemoglobins are produced. The unique globin structures of embryonic hemoglobin allow for adequate oxygen exchange in the relatively low-oxygen environment of early fetal life. 

As the fetal circulation develops and oxygen is obtained from the placenta (which provides higher oxygen concentrations than the yolk sac, but still lower than will eventually be provided by the lungs), another form of hemoglobin, called fetal hemoglobin, appears. Fetal hemoglobin persists throughout the rest of gestation, and is gradually replaced by adult hemoglobin during the first few months following birth. 

Finally, adult hemoglobin, which is predominant by six months after birth, is optimized for oxygen exchange between the high-oxygen environment of the lungs and the low-oxygen environment of the peripheral tissues.

These three normal human hemoglobins are characterized by different globins. Normal adult hemoglobin (called hemoglobin A) consists of two alpha and two beta globins. Hemoglobin A2 consists of two alpha and two delta globins. Fetal hemoglobin (hemoglobin F) contains two alpha and two gamma (beta-like) globins. There are various types of embryonic hemoglobin that contain several combinations of alpha, gamma, zeta, and epsilon globins.

Hemoglobinopathies

Numerous genetic mutations have been discovered that result in abnormalities of either the alpha-like or beta-like globins of the hemoglobin molecule. The abnormal hemoglobins resulting from these mutations are called hemoglobinopathies.

Over 1,000 kinds of hemoglobinopathies have been characterized so far. The majority of these are of minor significance and do not appear to cause clinical problems. They have been discovered, largely incidentally, in apparently normal people with the advent of screening hemoglobin electrophoresis tests.

However, several hemoglobinopathies do produce disease. The severity of a hemoglobinopathy usually depends on whether the mutation is homozygous (inherited from both parents), or heterozygous (inherited from only one parent, with normal hemoglobin genes from the second parent). In general, with heterozygous hemoglobinopathies, enough "normal" hemoglobin is produced to mitigate to at least some degree any overall clinical manifestations. People with homozygous forms of hemoglobinopathy tend to have more severe clinical disease.

The hemoglobinopathies are generally divided into two categories: 

• Hemoglobinopathies that are manifested by structural or functional changes in the hemoglobin molecule. These are generally referred to as the structural hemoglobinopathies.
• Hemoglobinopathies that are manifested by reduced production of one of the globin chains. Hemoglobinopathies in this second category are called the thalassemias.

Structural Hemoglobinopathies

Several structural hemoglobinopathies have been identified that produce clinical disease. Structural changes in the hemoglobin molecule can cause alterations in the shape and the flexibility of the red blood cells. The misshapen red cells can cause occlusion of blood vessels. Other kinds of structural hemoglobinopathies can cause hemolytic anemia. Yet other structural abnormalities can change the affinity of hemoglobin for oxygen.

Common structural hemoglobinopathies include:

• Hemoglobin S (sickle cell) is caused by a mutation in the beta globin chain. The homozygous form of sickle cell disease can cause the occlusion of blood vessels during periods of stress (sickle cell crisis), leading to pain, infections, swelling, and even more dire consequences such as stroke. The heterozygous form is called sickle cell trait and usually does not cause clinical problems.
• Hemoglobin C, hemoglobin D, and hemoglobin E are all caused by mutations in the beta globin chain. The homozygous forms of these disorders produce hemolytic anemia and spleen enlargement. The heterozygous forms usually do not produce significant disease.

Reduced Hemoglobin Production

The thalassemias are hemoglobinopathies that are caused by abnormalities in the genes that control the production of globins. This causes a reduced production of one of the globin chains, resulting in reduced production of hemoglobin, and thus anemia. People with thalassemia can also suffer from iron overload and an increased risk of infections.

Alpha thalassemias, most commonly seen in people of Asian or African descent, result in reduced alpha globin production. Beta thalassemias, most often seen in people of Mediterranean descent, cause reduced production of beta globin.

The thalassemias are genetically complex disorders, as several genetic mutations (alone or in combination) can produce thalassemia. The severity of thalassemia depends upon which globin chain is involved, and how many and which specific genes are causing the problem.

Combination Hemoglobinopathies

Occasionally, people inherit different hemoglobinopathy genes from each parent, resulting in what is called a compound heterozygous hemoglobinopathy or combination hemoglobinopathy. The more common combination hemoglobinopathies include:

• Hemoglobin SC disease, in which hemoglobin S comes from one parent and hemoglobin C comes from the other. Clinically, people with hemoglobin SC tend to have a milder form of sickle cell disease, but manifestations can vary widely.
• Sickle/beta-thalassemia, in which hemoglobin S comes from one parent and beta-thalassemia comes from the other. These people can have typical manifestations of sickle cell disease and anemia.

Follow-Up

Once the hemoglobinopathy has been fully characterized, you should expect your healthcare provider to have a detailed discussion with you over two topics: treatment you may need (if any), and genetic counseling.

If your hemoglobinopathy is the heterozygous form (so-called hemoglobin "trait," in which you have inherited the abnormal hemoglobin from only one parent), between 45% to 65% of your hemoglobin very likely will be normal adult hemoglobin, and your symptoms, if any, are likely to be mild. Most people with hemoglobin traits do not require any specific treatment.

If you have a homozygous hemoglobinopathy, or a combination hemoglobinopathy (that is, two different abnormal hemoglobins), you may need treatment. 

People with sickle cell disease today are almost always diagnosed in infancy with routine hemoglobin screening tests. These babies are treated with antibiotic prophylaxis, vitamin supplementation, full vaccination, and aggressive management of a sickle cell crisis whenever it occurs.

The thalassemias are a group of disorders whose effects vary widely depending on the specific genetic mutation that causes them. The most common problem they cause is anemia, but thalassemia can also cause skeletal abnormalities and iron overload as well as growth impairment and other disorders. People with severe thalassemia may require frequent blood transfusions and splenectomy. Iron overload can become a major problem in people with thalassemia.

Several uncommon hemoglobinopathies lead to "unstable hemoglobins," where the structure of the hemoglobin molecules is altered in such a way to reduce the lifespan of the red blood cells. People with these conditions may experience anemia, enlarged spleens, and frequent infections. Treatment is aimed at preventing complications and may include blood transfusions, splenectomy, and avoiding oxidant drugs, including certain antibiotics and NSAIDs. Bone marrow transplantation is also being applied more often to people with severe, life-threatening hemoglobinopathies.

Genetic Counseling

If the risk of producing a baby with a serious hemoglobinopathy is judged to be elevated, fetal evaluation may be indicated when pregnancy occurs.