Lab Testing and Hemoglobin Abnormalities

Hemoglobin abnormalities are variant forms of hemoglobin that are frequently inherited and can cause hemoglobinopathy (a blood disorder). 

Hemoglobin is a protein compound that contains iron and is found inside red blood cells. It transports oxygen throughout the entire body. It is comprised of globin chains, which are the proteins and heme, which is the part that contains iron.

There are several different kinds of globin chains: gamma, delta, and alpha. Regular types of hemoglobin include the following:  

Hemoglobin F (fetal hemoglobin or Hb F): Around 1% to 2% of hemoglobin that is found in adults. It has two gamma protein chains and two alpha protein chains. This is the main hemoglobin that a fetus produces during pregnancy. Usually, its product drops right after birth, and within 1-2 years reaches the adult level.  

Hemoglobin A2 (Hb A2): Around 2-3% of the hemoglobin that is found in adults. It contains two delta and two alpha protein chains. 

Hemoglobin A (Hb A): Around 95% to 98% of hemoglobin that is found in adults. Hemoglobin A contains two beta and two alpha protein chains.  

Mutations (genetic changes) within the globin genes result in globin protein alterations, which result in structurally altered hemoglobin, like hemoglobin S, which can cause thalassemia (reduction in global chain production) or sickle cell. With thalassemia, when the production of a globin chain is reduced, it upsets the balance of the beta and alpha chains and causes the formation of abnormal hemoglobin (alpha-thalassemia), or it can cause an increase in minor components of hemoglobin, such as Hb F (beta-thalassemia) or Hb A2.  

There are two genes each that code for gamma, delta, and beta globin chains and four that code for alpha globin chains. Mutations can occur in either the beta or alpha globin genes. The alpha thalassemia is the most common type of alpha-chain-related condition. The severity of the condition will depend on how many genes have been affected.  

Beta gene mutations are mainly inherited in an autosomal recessive manner. That means the individual must have two copies of altered genes, one from each of their parents, to have a hemoglobin variant disease. If one abnormal beta gene and one normal beta gene are inherited, the individual is heterozygous for abnormal hemoglobin, which is referred to as a carrier. The person’s abnormal gene may be passed onto children. However, it usually does not cause the carrier any significant health concerns or symptoms.  

If two of the same type of abnormal beta genes are inherited, then the individual is homozygous. The associated hemoglobin variant will be produced by the person, and they might potentially have some associated symptoms and complications. How severe the condition is will depend on the specific genetic mutation and will vary from one individual to the next. A copy of their abnormal beta gene is passed onto any children.  

If a person inherits two different types of abnormal beta genes, then the individual is “compound heterozygous” or “doubly heterozygous.” The affected individual typically will have symptoms that are related to both or one of the hemoglobin variants that the person produces. One abnormal beta gene will pass onto any children.  

Red blood cells that contain abnormal hemoglobin might not efficiently carry oxygen and might be broken down soon by the body than normal (shortened survival), which results in hemolytic anemia. There have been several hundred variants of hemoglobin documented. However, just a few of them are clinically significant and common. Some of the more common variants of hemoglobin include hemoglobin E, which can cause generally mild or no symptoms; hemoglobin C, which might cause minor hemolytic anemia; and hemoglobin S, which is the main hemoglobin in individuals who have sickle cell disease that can cause red blood cells to turn misshapen (sickle), which reduces the cells’ survival.  

Common Hemoglobin Variants 

There have been several hundred different hemoglobin variants (abnormal forms) that have been identified. However, just a few of them are clinically significant and common.  

E: this is one of the world’s most common types of beta chain hemoglobin variants. In Southeast Asia, it is especially prevalent, particularly in Thailand, Laos, and Cambodia, and in people of Southeast Asian descent. In individuals homozygous for Hb E (have two beta chain copies), usually have a mildly enlarged spleen, microcytic red blood cells, and mild hemolytic anemia. One hemoglobin E gene copy will not cause any symptoms unless another mutation combines with it, like the beta-thalassemia trait one.  

C: Around 2-3% of U.S. African Americans are heterozygous for hemoglobin C (one copy called hemoglobin C trait). They are frequently asymptomatic. The hemoglobin C disease (those with two copies, seen in homozygotes) is relatively mild and rare (0.02% of U.S. African Americans). Usually, it causes a mild or moderately enlarged spleen and minor hemolytic anemia.  

S: this is the main hemoglobin in individuals who have sickle cell disease (which is also referred to as sickle cell anemia). The Centers for Disease Control and Prevention reports that an estimated 1 in 375 African American infants are born with sickle cell anemia, and around 100,000 Americas have this disorder. People who have Hb S disease possess two normal alpha chains and two abnormal beta chains. Hb S causes red blood cells to become deformed and turn into the sickle shape when they are exposed to reduced amounts of oxygen (like what may occur when someone has a lung infection or exercises). Sickle red blood cells are very rigid and may result in small blood vessels becoming blocked, which decreases oxygen delivery, impair circulation, cause pain, and shorten the survival of red blood cells. One beta copy (called the sickle cell trait and present in an estimated 8% of African Americans) usually doesn’t cause any serious symptoms unless another hemoglobin mutation combines with it, like that which causes beta-thalassemia or Hb C. 

Less Common Variants   

Many other variants exist. Some of them are silent and cause no symptoms or signs. Then others might affect the stability and/or functionality of the hemoglobin molecules. Other variants include Hb M, Hb J, Hb G, Hb D, and Hb Constant Spring, which is caused by a mutation within the alpha globin gene that causes an unstable hemoglobin molecule and abnormally long alpha chain.

Other examples include the following:  

F: Hb F is the main hemoglobin that the fetus produces. Its role is to efficiently transport oxygen within a low oxygen environment. Hb F production is sharply reduced after birth and by 1-2 years old reaches adult levels. In several different congenital disorders, Hb F might be elevated. In beta-thalassemia, levels can be significantly increased or normal, and in sickle cell beta-thalassemia or people with sickle cell anemia, levels are frequently increased. People with increased Hb F and sickle cell disease frequently have a milder form of the disease, since the sickling of red cells is inhibited by the F hemoglobin. Hb F levels also are increased in the hereditary persistence of fetal hemoglobin (HPFH), which is a rare condition. In these inherited disorders, there are increased Hb F levels without the clinical features or signs of thalassemia. Various ethnic groups have various mutations that may cause HPFH. Also, Hb F may be increased as well in certain acquired conditions that involved the impaired production of red blood cells. Some leukemias, as well as other types of myeloproliferative neoplasms, also are associated with mild increases in Hb F.  

H: The abnormal hemoglobin Hb H occurs in some alpha thalassemia cases. It is comprised of four beta globin chains. It is produced due to a serious shortage in alpha chains. The beta globin chains are all normal. However, on 4 of the beta chains, the tetramer does not function properly. Its affinity for oxygen is increased and holds onto it rather than releasing it to the cells and tissues. Hemoglobin H also is associated with hemolysis (serious breakdown in red blood cells) since it is very unstable and tends to form solid structures inside the red blood cells. Individuals who have hemoglobin H disease often have anemia but do not usually have serious medical problems.  

Barts: This develops in fetuses that have alpha thalassemia. Hb Barts is formed from four gamma protein chains whenever there are not enough alpha chains, in a way that is like Hemoglobin H formation. If small Hb Bart levels are detected, normally it disappears soon after birth because gamma chain production dwindles. These children are silent carriers with one or two deletions of alpha genes or possess the alpha thalassemia trait. A child with large Hb Barts levels usually will have a three-gene deletion and hemoglobin H disease. Fetuses that have four-gene deletions will have hydrops fetalis and normally will not survive without bone marrow transplants and blood transfusions.   

Two separate abnormal genes can also be inherited by a person, one from each of their parents. It is referred to as being doubly heterozygous or compound heterozygous. Below are listed several different combinations that are clinically significant. 

SC disease: inheriting one beta C gene and one beta S gene causes Hemoglobin SC disease. These people have moderately enlarged spleens and mild hemolytic anemia. Individuals with Hb SC disease might develop the same blood vessel-blocking (vaso-occlusive) complications that are found in sickle cell anemia. However, most cases are not as serious.  

D disease Sickle Cell: People with sickle cell or Hb D disease inherit one copy of hemoglobin D and one hemoglobin S. Those individuals might have moderate hemolytic anemia and occasional sickle crises.  

E-beta thalassemia: People who are doubly heterozygous for beta thalassemia and hemoglobin E have anemia that may vary in severity, ranging from mild (asymptomatic) up to severe, depending on what beta thalassemia mutation(s) are present.  

S-beta thalassemia: Beta thalassemia – sickle cell various in severity, which depends on the inherited beta thalassemia mutation. Some mutations can result in the reduced production of beta globin (beta+), where it is eliminated (beto0) by others. Sickle cell beta+ thalassemia tends to be less severe compared to beta0 thalassemia. Individuals with sickle cell – beta thalassemia tend to have an increased number of irreversibly sickled cells, more serious anemia, and more frequent vaso-occlusive issues compared to people who have sickle cell – beta thalassemia. Often it is hard to distinguish between sickle cell – beta thalassemia and sickle cell disease.  

Symptoms and Signs  

Symptoms and signs that are associated with hemoglobin variations vary in severity and type depending on which variant is present and whether the person has a combination or one variant. Some are due to an increase in hemolysis (breakdown) of red blood cells as well as shortened red blood cell survival, which results in anemia.

The following are some examples:  

  • Pale skin (pallor) 
  • Jaundice 
  • Lack of energy 
  • Weakness, fatigue 
  • Some serious symptoms and signs include:  
  • Upper abdomen pain (caused by the formation of stones in the gallbladder) 
  • Growth problems for children 
  • Enlarged spleen 
  • Shortness of breath  
  • Severe pain episodes  

Laboratory Tests 

Hemoglobin variant lab tests explore the “normalness” of a person’s red blood cells, analyze relevant gene mutations, and/or evaluate the hemoglobin with the red blood cells. Each test offers a piece of the overall puzzle to provide important information to the clinician about whatever hemoglobin might be present.

Typically testing includes:  

Complete blood count (CBC): This test provides a snapshot of the cells that are circulating within the blood. The CBC, among other things, will let the doctor know the number of red blood cells that are present, the amount of hemoglobin inside of them, and provide an evaluation to the doctor of red blood cells’ average size.  

Mean corpuscular volume (MCV) measures the red blood cells’ size. Low MCV frequent is an early indication of thalassemia. If there is low MCV and iron-deficiency is ruled out, then the person might have a hemoglobin variant that is the result of red blood cells that are smaller than normal (Hb E, for example) or be a thalassemia trait carrier.  

Blood smear (or peripheral smear): A trained laboratorian will look under a microscope at a thin blood layer on a slide that has been treated using a special stain. The type and number of platelets, red blood cells, and white blood cells can be evaluated in order to determine whether they are mature and normal.

With hemoglobinopathy, red blood cells might be:  

  • Microcytic (smaller than normal) 
  • Hypochromic (paler than normal)  
  • Vary in shape (poikilocytosis – e.g., sickle-shaped cells) and size (anisocytosis) 
  • Have a crystal (e.g., C crystal) or nucleus (nucleated red blood cell, which in mature red blood cells is not normal) 
  • Uneven distribution of hemoglobin (“target cells” are produced that under a microscope resemble a bull’s eye).  

The higher the percentage of abnormal-appearing red blood cells there are, the higher the chance that an underlying disorder is present.  

Hemoglobinopathy evaluation: This type of test identifies the type and measures the relative amount of the various kinds of hemoglobin that are present in a person’s red blood cells. Most common variants may be identified using a combination or one of the tests. The relative amount of variant hemoglobin that is detected can help with diagnosing combinations of thalassemia (compound heterozygotes) and hemoglobin variants.  

Genetic testing: This type of test is used for investigating the mutations and deletions in the beta and alpha globin-producing genes. It is possible to conduct family studies to evaluate both the carrier status as well as the kinds of mutations that are present in other members of the family. Genetic testing is not done regularly. However, it may be used to help determine carrier status and confirm thalassemia and hemoglobin variants. 

Laboratory Tests 

Sickle Cell Tests 

Hemoglobinopathy Evaluation 

Hemoglobin 

Iron-binding and Transferrin Capacity (UIBC, TIBC) 

Iron 

Ferritin 

Blood Smear 

Complete Blood Count  

Conditions 

Pregnancy: Preconception 

Thalassemia 

Sickle Cell Anemia 

Anemia