Complete blood count
A complete blood count (CBC) is a series of tests used to evaluate the composition and concentration of the cellular components of blood. It consists of the following tests: red blood cell (RBC) count, white blood cell (WBC) count, and platelet count; measurement of hemoglobin and mean red cell volume; classification of white blood cells (WBC differential); and calculation of hematocrit and red blood cell indices . The hematocrit is the percentage of blood by volume that is occupied by the red cells (i.e., the packed red cell volume). Red blood cell indices are calculations derived from the red blood cell count, hemoglobin, and hematocrit that aid in the diagnosis and classification of anemia.
The CBC provides valuable information about the blood and to some extent the bone marrow, which is the blood-forming tissue. The CBC is used for the following purposes:
- as a preoperative test to ensure both adequate oxygen carrying capacity and hemostasis
- to identify persons who may have an infection
- to diagnose anemia
- to identify acute and chronic illness, bleeding tendencies, and white blood cell disorders such as leukemia
- to monitor treatment for anemia and other blood diseases
- to determine the effects of chemotherapy and radiation therapy on blood cell production
The CBC requires a sample of blood collected from a vein. The nurse or phlebotomist inserting the needle should clean the skin first. The tourniquet should be removed from the arm as soon as the blood flows. If a fingerstick is used to collect the blood, care must be taken to wipe away the first drop, and not to squeeze the finger excessively as this causes the blood to be diluted by tissue fluid. Many drugs affect the results by causing increased or decreased RBC, WBC, and/or platelet production. Medications should be taken into account when interpreting results.
The CBC is commonly performed on an automated hematology analyzer using well mixed whole blood that is added to a chemical called EDTA to prevent clotting. A CBC is a group of tests used to quantify the number of RBCs, WBCs, and platelets, provide information about their size and shape, measure the hemoblobin content of RBCs, determine the percentage and absolute number of the five white blood cell types, and identify early and abnormal blood cells. These tests are performed simultaneously, (usually in less than one minute), using an automated hematology analyzer. When the performance limit of the automated hematology analyzer is exceeded, sample dilution or pretreatment, manual smear review, or manual cell counts may be required. Each laboratory has established rules for determining the need for manual smear review based upon specific CBC parameters. For example, a manual differential is always performed when nucleated immature red blood cells are found on an electronic cell count.
Electronic cell counting
Electronic blood cell counting is based upon the principle of impedance (i.e., resistance to current flow). Some hematology analyzers combine impedance counting with light scattering to measure platelets. A small sample of the blood is aspirated into a chamber (the WBC counting bath) and diluted with a balanced isotonic saline solution that is free of particles. The diluted blood sample is split into two parts, one for counting RBCs and platelets and the other for counting WBCs. The RBC portion is transferred to the RBC/platelet counting bath where it is diluted further. The other portion remains in the WBC bath and a detergent (lysing agent) is added to destroy (hemolyze) the red blood cells. A small portion of the diluted fluid in each bath is allowed to flow past a small aperture. An electrical current is produced in each aperture by two electrodes, one on the inside and the other on the outside of the aperture. The saline solution is responsible for conducting current between the electrodes. The cells move through the aperture one at a time. When a cell enters the aperture, it displaces a volume of electrolyte equal to its size. The cell acts as an electrical resistor, and impedes the flow of current. This produces a voltage pulse, the magnitude of which is proportional to the size of the cell. Instrument electronics are adjusted to discriminate voltage pulses produced by different cells. These adjustments are called thresholds. For example, the threshold for counting a RBC is equivalent to a cell volume of 36 femtoliters or higher. Voltage pulses that are equivalent to volumes of 2–20 femtoliters are counted as platelets. This process is repeated two more times so that the RBC, WBC, and platelet counts are performed in triplicate. Each time frame for counting is several seconds and many thousands of cells are counted. The computer processes the counting data first by determining the agreement between the three counts. If acceptable criteria are met, the counts are accepted and used to calculate the result.
The hemoglobin concentration is measured optically using the solution in the WBC bath. The lysing agent contains potassium cyanide that reacts with the hemoglobin to form cyanmethemoglobin. The optical density of the cyanmethemoglobin is proportional to hemoglobin concentration.
The voltage pulses produced by the white blood cells depend upon the size of the cell and its nuclear density. Therefore, the pulses may be analyzed to differentiate between the types of WBCs found. For example, lymphocytes are the smallest WBCs and comprise the lower end of the size scale. Monocytes, prolymphocytes, and immature granulocytes comprise the central area of the WBC histogram, and mature granulocytes comprise the upper end. In addition to cell sizing, automated instruments may use any of three other methods to distinguish between subpopulations. These are radio frequency conductance, forward and angular light scattering, and fluorescent staining.
Red blood cell count
The red cells, the most numerous of the cellular elements, carry oxygen from the lungs to the body’s tissues. They are released from the bone marrow into the blood in an immature form called the reticulocyte that still retains much of the cellular RNA needed for hemoglobin production. Reticulocytes may be counted on some automated analyzers and are an index to recovery from anemia. The average life span of RBCs in the circulation is approximately 120 days.
The red blood cell (RBC) count determines the total number of red cells (erythrocytes) in a sample of blood. Most anemias are associated with a low RBC count, hemoglobin, and hematocrit. Common causes include excessive bleeding; a deficiency of iron, vitamin B 12 , or folic acid; destruction of red cells by antibodies or mechanical trauma; bone marrow malignancy and fibrosis; and structurally abnormal hemoglobin. The RBC count is also decreased due to cancer, kidney diseases, and excessive IV fluids. An elevated RBC count may be caused by dehydration, hypoxia (decreased oxygen), or a disease called polycythemia vera. Hypoxia may result from high altitudes, chronic obstructive lung diseases, and congestive heart failure.
Hematocrit and cell indices
The hematocrit is a test that measures the volume of blood in percent that is comprised of the red blood cells. Automated cell counters calculate the hematocrit by multiplying the RBC count by the mean red cell volume. A decrease in the number or size of red cells also decreases the amount of space they occupy, resulting in a lower hematocrit. Conversely, an increase in the number or size of red cells increases the amount of space they occupy, resulting in a higher hematocrit. Thalassemia minor, a genetic cause of anemia, is an exception in that it usually causes an increase in the number of red blood cells, but because they are small, it results in a decreased hematocrit.
The three main RBC indices are used to determine the average size and hemoglobin content of the RBCs and they help determine the cause of anemia. The three indices are described below:
- Mean corpuscular volume (MCV)—the average size of the red blood cells expressed in femtoliters. MCV is calculated by dividing the hematocrit (as percent) by the RBC count in millions per microliter of blood, then multiplying by 10.
- Mean corpuscular hemoglobin (MCH)—the average amount of hemoglobin inside an RBC expressed in picograms. The MCH is calculated by dividing the hemoglobin concentration in grams per deciliter by the RBC count in millions per microliter, then multiplying by 10.
- Mean corpuscular hemoglobin concentration (MCHC)—the average concentration of hemoglobin in the RBCs expressed in percent. It is calculated by dividing the hemoglobin in grams per deciliter by the hematocrit, then multiplying by 100.
The mechanisms by which anemia occurs will alter the RBC indices in a predictable manner. Therefore, the RBC indices permit the physician to narrow down the possible causes of an anemia. The MCV is an index of the size of the RBCs. When the MCV is below normal, the RBCs will be smaller than normal and are described as microcytic. When the MCV is elevated, the RBCs will be larger than normal and are termed macrocytic. RBCs of normal size are termed normocytic. Failure to produce hemoglobin results in smaller than normal cells. This occurs in many diseases including iron deficiency anemia, thalassemia (an inherited disease in which globin chain production is deficient), and anemias associated with chronic infection or disease. Macrocytic cells occur when division of RBC precursor cells in the bone marrow is impaired. The most common causes of macrocytic anemia are vitamin B 12 deficiency, folate deficiency, and liver disease. Normocytic anemia may be caused by decreased production (e.g., malignancy and other causes of bone marrow failure), increased destruction (hemolytic anemia), or blood loss. The RBC count is low, but the size and amount of hemoglobin in the cells is normal.
White blood cell count
The majority of CBCs include both a WBC count and an automated differential. A differential determines the percentage of each of the five types of mature white blood cells. An elevated WBC count occurs in infection, allergy, systemic illness, inflammation, tissue injury, and leukemia. A low WBC count may occur in some viral infections, immunodeficiency states, and bone marrow failure. The WBC count provides clues about certain illnesses, and helps physicians monitor a patient’s recovery from others. The differential will reveal which WBC types are affected most. For example, an elevated WBC count with an absolute increase in lymphocytes having an atypical appearance is most often caused by infectious mononucleosis. The differential will also identify early WBCs that may be reactive (e.g., a response to acute infection) or the result of a leukemia.
When the electronic WBC count is abnormal or a cell population is flagged, meaning that one or more of the results is atypical, a manual differential is performed. In that case, a wedge smear is prepared. This is done by placing a drop of blood on a glass slide, and using a second slide to pull the blood over the first slide’s surface. The smear is air dried, then stained with Wright stain and examined under a microscope using oil immersion (1000x magnification). One hundred white cells are counted and identified as either neutrophils, lymphocytes, monocytes, eosinophils, or basophils based on the shape and appearance of the nucleus, the color of cytoplasm, and the presence and color of granules. The purpose is to determine if these cells are present in a normal distribution, or if one cell type is increased or decreased. Any atypical or immature cells also are counted.
In addition to determining the percentage of each mature white blood cell, the following tests are performed as part of the differential:
- Evaluation of RBC morphology is performed. This includes grading of the variation in RBC size (anisocytosis) and shape (poikioocytosis); reporting the type and number of any abnormal RBCs such as target cells, sickle cells, stippled cells, etc.; reporting the presence of immature RBCs (polychromasia); and counting the number of nucleated RBCs per 100 WBCs.
- An estimate of the WBC count is made and compared to the automated or chamber WBC count. An estimate of the platelet count is made and compared to the automated or chamber platelet count. Abnormal platelets such as clumped platelets or excessively large platelets are noted on the report.
- Any immature white blood cells are included in the differential count of 100 cells, and any inclusions or abnormalities of the WBCs are reported.
WBCs consist of two main subpopulations, the mononuclear cells and the granulocytic cells. Mononuclear cells include lymphocytes and monocytes. Granulocytes include neutropohils (also called polymorphonuclear leukocytes or segmented neutrophils), eosinophils, and basophils. Each cell type is described below:
- Neutrophils are normally the most abundant WBCs. They measure 12–16 μm in diameter. The nucleus stains dark purple-blue, and is divided into several lobes (usually three or four) consisting of dense chromatin. A neutrophil just before the final stage of maturation will have an unsegmented nucleus in the shape of a band. These band neutrophils may be counted along with mature neutrophils or as a separate category. The cytoplasm of a neutrophil contains both primary (azurophilic) and secondary (specific) granules. The secondary granules are lilac in color and are more abundant, almost covering the pink cytoplasm. Neutrophils are phagocytic (able to engulf objects) cells and facilitate removal of bacteria and antibody-coated antigens. The neutrophilic granules are rich in peroxidase, and aid the cell in destroying bacteria and other ingested cells.
- Eosinophils are 14–16 μm in diameter and contain a blue nucleus that is segmented into two distinct lobes. The cytoplasm is filled with large refractile orange-red granules. The granules contain peroxidase, hydrolases, and basic proteins that aid in the destruction of phagocytized cells. Eosinophils are increased in allergic reactions and parasitic infections.
- Basophils, like eosinophils, are 14–16 μm in diameter and have a blue nucleus that is bilobed. The cytoplasm of the basophil is filled with large dark blue-black granules that may obscure the nucleus. These contain large amounts of histamine, heparin, and acid mucopolysaccharides. Basophils mediate the allergic response by releasing histamine.
- Lymphocytes are the second most abundant WBCs. They may be small (7–9 μm in diameter) or large (12–16 μm in diameter). The nucleus is dark blue and is nearly round or slightly indented and the chromatin is clumped and very dense. The cytoplasm is medium blue and usually agranular. An occasional lymphocyte will have a few azurophilic granules in the cytoplasm. Lymphocytes originate in the lymphoid tissues and are not phagocytic. They are responsible for initiating and regulating the immune response by the production of antibodies and cytokines.
- Monocytes are the largest WBCs, measuring 14–20 μm in diameter. They have a large irregularly shaped and folded blue nucleus with chromatin that is less dense than other WBCs. The cytoplasm is gray-blue, and is filled with fine dust-like lilac colored granules. Monocytes are phagocytic cells that process and present antigens to lymphocytes, an event required for lymphocyte activation.
Platelets are disk-shaped structures formed by the detachment of cytoplasm from megakaryocytes. They aid in the coagulation process by attaching or adhering to the walls of injured blood vessels, where they stick together to form the initial platelet plug. A low platelet count may occur in patients with AIDS, viral infections, lymphoma, and lupus erythematosus, or in patients taking certain drugs, most notably quinine and quinidine. Decreased platelet production is also a cause of thrombocytopenia, and may be due to aplastic anemia, leukemia, lymphoma, or bone marrow fibrosis. A low platelet count can occur due to increased destruction. This can result from antibody production that is often drug-induced (heparin treatment being a prominent cause). Increased destruction also results from autoantibody production as occurs in idiopathic thrombocytopenic purpura (ITP) and thrombotic episodes that consume platelets such as occur in thrombotic thrombocytopenic purpura (TTP), disseminated intravascular coagulation (DIC), and hemolytic-uremic syndrome (HUS). Inherited (congenital) thrombocytopenia can be caused by Glanzmann’s thrombasthenia, Fanconi syndrome, and Wiskott-Aldrich syndrome.
Thrombocytosis, an increased platelet count, is most often caused by a reaction to injury or inflammation. In these cases the platelet count increases transiently and is usually within the range of 400,000–800,000 per microliter. Persistent or higher counts are usually associated with myeloproliferative disease (malignant disease involving blood forming cells) such as chronic granulocytic (myelogenous) leukemia, polycythemia vera, or primary (essential) thrombocythemia.
The platelet count is most often measured by impedance counting but is performed manually when the platelet count is very low, platelet clumping is observed, or abnormally large (giant) platelets are present. Often these abnormalities and others such as cryoglobulinemia, cell fragmentation (hemolysis), and microcytic RBCs are signaled by abnormal RBC and platelet indices and abnormal population flags. An abnormal mean platelet volume or platelet histogram indicates that morphological platelet abnormalities are present and the platelets should be observed from a stained blood film to characterize the abnormality. The platelet count can be estimated using the Wright-stained blood smear used for a differential WBC count by multiplying the average number of platelets per oil immersion field by 20,000. Platelet estimates should correlate with actual counts. When they disagree, the platelet count should be repeated and a manual count performed if necessary.
The CBC does not require fasting or any special preparation.
Discomfort or bruising may occur at the puncture site. Applying pressure to the puncture site until the bleeding stops helps to reduce bruising; warm packs relieve discomfort. Some people feel dizzy or faint after blood has been drawn and should be treated by resting awhile.
Other than potential bruising at the puncture site, and/or dizziness, there are no complications associated with this test.
CBC values vary by age and sex. Normal values are ultimately determined by the laboratory performing the test. As a guide, the normal values for men and nonpregnant women are as follows:
- WBCs: 4,500–11,000 per microliter for women and men, with neutrophils representing 50–70%, lymphocytes 25–35%, monocytes 4–6%, eosinophils 1–3%, basophils 0.4–1%, and bands 0–5%.
- RBCs: 4.2–5.0 million per microliter for women; 4.5–6.2 million per microliter for men.
- Hemoglobin: 12–15 g/dL for women; 13.6–17.2 g/dL for men.
- Hematocrit: 35–47% for women; 42–52% for men.
- Platelets: 150,000 and 350,000 per microliter.
- Reticulocyte count: 0.5–1.5%.
Normal adult results for red blood cell indices are as follows:
- MCV: 80–98 fl (femtoliters)
- MCHC: 32–36%
- MCH: 27–31 pg (picograms)
- RDW: 11.5–14.5%
In addition to normal values, critical values (alert, panic values) are established for hemoglobin (and hematocrit), WBC count, and platelet count. Precipitously low hemoglobin is associated with hypoxia that can have life-threatening complications. Extremely low WBCs indicates an inability to fight infection and a high risk of sepsis. A severely reduced platelet count predisposes the patient to spontaneous internal bleeding. Representative critical values are shown below.
- Hemoglobin: less than 5.0 g/dL
- Hematocrit: less than 15%
- Platelet count: less than 30,000 per microliter
- WBC count: less than 2,500 per microliter and greater than 30,000 per microliter
Abnormal blood count results are seen in a variety of conditions. One of the most common is anemia, which is characterized by a low RBC count, hemoglobin, and hematocrit. The category into which a person’s anemia is placed is in part based upon the red blood cell indices provided. The indices provide a significant clue as to the cause of the anemia, but further testing is needed to confirm a specific diagnosis. The most common causes of macrocytic anemia (high MCV) are vitamin B 12 and folic acid deficiencies. Lack of iron in the diet, thalassemia (a type of hereditary anemia), and chronic illness are the most common causes of microcytic anemia (low MCV). Normocytic anemia (normal MCV) can be caused by kidney and liver disease, bone marrow disorders, leukemia, excessive bleeding, or hemolysis of the red blood cells. Iron deficiency and thalassemia are the most common causes of hypochromic anemia (low MCHC). Normocytic anemias are usually also normochromic and share the same causes. The red cell distribution width (RDW) is increased in anemias caused by deficiencies of iron, vitamin B 12 , or folic acid. Abnormal hemoglobins, such as in sickle cell anemia, can change the shape of red blood cells as well as cause them to hemolyze, or rupture. The abnormal shape and the cell fragments resulting from hemolysis increase the RDW. Conditions that cause more immature cells to be released into the bloodstream, such as severe blood loss, will increase the RDW. The larger size of immature cells creates a distinct size variation.
Infections and leukemias are associated with increased numbers of WBCs. Increases or decreases in the percentage of each white cell can be associated with a number of diseases or conditions, including cancer, leukemia, anemia, multiple sclerosis, allergies, parasitic and viral diseases, infections, and tissue damage