Clinical Significance of Anemia Tests

Active-B12 (AB12)
The Active-B12 assay is typically used as an aid in the diagnosis and treatment of vitamin B12 deficiency.8 The Active-B12 assay measures holotranscobalamin (holoTC) in the blood. HoloTC is a result of the B12 vitamin (cobalamin) binding with the protein transcobalamin, which transports vitamin B12. HoloTC is also referred to as Active-B12 because it is the only form of vitamin B12 that can be taken up by the cells of the body. 

Causes of vitamin B12 deficiency include not enough vitamin B12 in the diet, diseases that cause malabsorption (Crohn’s disease), lack of intrinsic factor (a protein that helps the intestine absorb vitamin B12), above-normal heat production, and pregnancy.7

Erythropoietin (EPO)

The erythropoietin test measures the amount of a hormone called erythropoietin (EPO) in blood. This test may be used to help determine the cause of anemia, polycythemia (high red blood cell count), or other bone marrow disorders.

A change in red blood cells affects the release of EPO. For example, people with anemia have too few red blood cells, so more EPO is produced. Increased EPO levels may be due to secondary polycythemia, an overproduction of red blood cells that occurs in response to an event such as low blood-oxygen levels. This may happen at high altitudes or, rarely, because of a tumor that releases EPO.3  Lower-than-normal EPO levels may be seen in chronic kidney failure, anemia of chronic disease, or polycythemia vera.3


Measurement of ferritin aids in the diagnosis of diseases affecting iron metabolism, such as hemochromatosis (iron overload) and iron-deficiency anemia. The ferritin test is typically ordered to assess iron stores in the body. The test is sometimes ordered along with an iron test and a TIBC to detect the presence and evaluate the severity of an iron deficiency or overload.4 

Early iron deficiency usually causes no physical effects at all. If a person is otherwise healthy, symptoms seldom appear before the hemoglobin in the blood drops below a certain level (10 g per deciliter). The ferritin test may be ordered, along with other iron tests, when a routine CBC shows that a person's hemoglobin and hematocrit are low and his or her red blood cells are smaller and paler than normal (microcytic and hypochromic). These conditions suggest iron-deficiency anemia, even though other clinical symptoms have not yet developed.4


Folate, along with vitamin B12, is primarily ordered to detect deficiencies and to help diagnose the cause of certain anemias. In those treated for known B12 and folate deficiencies, these tests will be ordered occasionally to monitor the effectiveness of treatment. This is especially true in those who cannot properly absorb B12 and/or folate and must have lifelong treatment.4 

Folate and B12 may also be ordered to aid in diagnosis when an individual presents with an altered mental state or other behavioral changes, especially in the elderly. B12 may be ordered with folate, by itself, or with other laboratory screening tests (antinuclear antibody, CRP, rheumatoid factor, CBC, and chemistry blood tests) to help establish reasons why a person shows symptoms of neuropathy.4 

Serum folate levels can vary based on dietary intake. RBCs can store 95% of circulating folate; thus a measurement of the RBC folate level may be used to help detect a deficiency. Some doctors believe that the RBC folate test is more clinically relevant than serum folate, but there is not widespread agreement on this.4


Low hemopexin levels are one of the diagnostic features of hemolytic anemia.5  Measurements of hemopexin are used as an aid in the diagnosis of various hemotologic disorders, such as hemolytic anemia and sickle cell anemia.


Homocysteine tests help determine if a person has B12 or folate deficiency. The homocysteine concentration may be elevated before B12 and folate tests are abnormal. Physicians may recommend homocysteine testing in malnourished patients and the elderly, who often absorb less vitamin B12 from their diets, and those with drug or alcohol addictions.4


Serum iron measures the amount of circulating iron that is bound to transferrin. Iron status may be evaluated by ordering one or more tests to determine the amount of iron in the blood, the capacity of the blood to transport iron, and the amount of iron in storage. The tests may also help differentiate various causes of anemia. These tests are often ordered together, and the results of each test can help the doctor to determine the cause of iron deficiency and/or overload.4 

Iron is also measured to check general nutritional status and to see if iron and nutritional treatment is working.

RBC Folate

Red cell folate (RBC folate) is a measure of the body’s store of the vitamin folate, also known as folic acid. A low RBC folate level means a deficiency of folate. Folate is important for the normal production of red blood cells, and in prevention of neural tube defects such as spina bifida, in the unborn babies of pregnant women.

An RBC folate test may be requested if a patient is suspected of having anemia, particularly of the macrocytic variety, where the red blood cells are increased in size.6

Soluble Transferrin Receptor (sTfR)

Soluble transferrin receptors are proteins found in blood that can be elevated with iron deficiency. The sTfR test measures the amount of soluble transferrin receptors to help detect and evaluate iron deficiency and aid in the diagnosis of iron-deficiency anemia.4 

sTfR is useful in detecting iron-deficiency anemia and distinguishing it from anemia caused by chronic illness or inflammation. Usually, a ferritin test is used to evaluate the amount of iron reserve in the body and to help diagnose iron-deficiency anemia. However, the ferritin level can be elevated when an individual has inflammation or a chronic disease, such as autoimmune disorders or some cancers. For these conditions, the ferritin test is not useful, and a test for soluble transferrin receptors may be ordered.4

Total Iron-binding Capacity (TIBC)

TIBC is generally measured to assess the body's ability to transport iron in the blood. TIBC measures all the proteins in the blood that are available to bind with iron, including transferrin. Since transferrin is the primary iron-binding protein, the TIBC test is a good indirect measurement of transferrin.4 The body produces transferrin in relationship to the need for iron. When iron stores are low, transferrin levels increase and vice versa. In healthy people, about one-third of the binding sites on transferrin are used to transport iron.

Vitamin B12

Vitamin B12 testing is typically used to diagnose the cause of anemia or neuropathy, to evaluate nutritional status, or to monitor the effectiveness of treatment for vitamin B12 or folate deficiency.4 

Vitamin B12 testing is most often done when other blood tests suggest a condition called megaloblastic anemia. Pernicious anemia is a form of megaloblastic anemia caused by poor vitamin B12 absorption. This can occur when the stomach makes less of the substance the body needs to properly absorb vitamin B12.7