HEMATOLOGY EBOOK

Red blood cell indices are blood tests that provide information about the hemoglobin content and size of red blood cells. Abnormal values indicate the presence of anemia and which type of anemia it is.

Mean corpuscular volume

The mean corpuscular volume, or “mean cell volume” (MCV), is a measure of the average red blood cell volume (i.e. size) that is reported as part of a standard complete blood count. In patients with anemia, it is the MCV measurement that allows classification as either a microcytic anemia (MCV below normal range) or macrocytic anemia (MCV above normal range).

Calculation

It can be calculated (in litres) by dividing the hematocrit by the red blood cell count (number of red blood cells per litre). The results is typically reported in femtolitres. If the MCV was determined by automated equipment, the result can be compared to RBC morphology on a peripheral blood smear. Any deviation would be indicative of either faulty equipment or technician error. For further specification, it can be used to calculate red blood cell distribution width (RDW).

Interpretation

The normal referance range is typically 80-100 fL.

High

In presence of hemolytic anaemia, presence of reticulocytes can increase MCV. In pernicious anemia (macrocytic), MCV can range up to 150 femtolitres. An elevated MCV is also asociated with alcoholism (as are an elevated GGT and a ratio of AST:ALT of 2:1). Vitamin b12 and/or Folic Acid deficiency has also been ssociated with macrocytic anemia (high MCV numbers).

Low

The most common causes of microcytic anemia are iron deficiency (due to inadequate dietary intake, gastrointestinal blood loss, or menstrual blood loss), thalassemia, or chronic disease. A low MCV number in a patient with a positivve stool guaiac test (bloody stool) is highly suggestive of GI cancer. In iron deficiency anemia (microcytic anemia), it can be as low as 60 to 70 femtolitres. In cases of thalassemia, the MCV may be low even though the patient is not iron deficient.

Mean corpuscular hemoglobin

The mean corpuscular hemoglobin, or “mean cell hemoglobin” (MCH), is the average of hemoglobin per red blood cell in a sample of blood. It is reported as part of a standard complete blood count: MCH value is diminished in hypochromic anemias. It is calculated by dividing the total mass of hemoglobin by the number of red blood cells in a volume of blood.

MCH=(Hgb*10)/RBC

A normal value in humans is 27 to 31 picograms/ cell. Conversion to SI-units: 1 pg of hemoglobin = 0,06207 femtomol. Normal value converted to SI-Units: 1,68 – 1,92 fmol/cell.

Mean corpuscular hemoglobin concentration

The mean corpuscular hemoglobin concentration, or MCHC, is a measure of the concentration of hemoglobin in a given volume of packed red blood cell. It is reported as part of a standard complete blood count. It is calculated by dividing the hemoglobin by the hematocrit. Reference ranges for blood tests are 32 to 36 g/dl, or between 4.9 to 5.5 mmol/L. It is thus a mass or molar concentration. Still, many instances measure MCHC in percentage (%), as if it was a mass fraction (m_Hb / m_RBC). Numerically, however, the MCHC in g/dl and the mass fraction of hemoglobin in red blood cells in % are identical, assuming a RBC density of 1g/mL and negligible hemoglobin in plasma.

Interpretation

It is diminished (“hypochromic”) in microcytic anemias, and normal (“normochromic”) in macrocytic anemias (due to larger cell size, though the hemoglobin amount or MCH is high, the concentration remains normal). MCHC is elevated in hereditary spherocytosis.

This count is used to give a rough guide to what shade of red, RBC will be. (paler=lower than the standard)

Complications with cold agglutinin

Because of the way automated analysers count blood cells, a very high MCHC (greater than about 370 g/L) may indicate the blood is from someone with a cold agglutinin. This means that when their blood gets colder than 37°C it starts to clump together. As a result, the analyzer may incorrectly report a low number of very dense red blood cells for blood samples in which agglutination has occurred.

This problem is usually picked up by the laboratory before the result is reported. The blood is warmed until the cells separate from each other, and quickly put through the machine while still warm.This is the most sensitive test for iron deficiency anemia.

Worked example

D-dimer is a fibrin degradation product, a small protein fragment present in the blood after a blood clot is degraded by fibrinolysis. It is so named because it contains two crosslinked D fragments of the fibrinogen protein. D-dimer concentration may be determined by a blood test to help to help diagnose thrombosis. Since its introduction in the 1990s, it has become an important test performed in patients suspected of thrombotic disorders. While a negative result practically rules out thrombosis, particularly in young and healthy patients, a positive result can indicate thrombosis but does not rule out other potential causes. Its main use, therefore, is to exclude thromboembolic disease where the probability is low. In addition, it is used in the diagnosis of the blood disorder disseminated intravascular coagulation

Principles

Principles of D-dimer testing

Coagulation, the formation of ablood clot of thrombin, occurs when the proteins of the “coagulation cascade” are activated, either by contact with damaged blood vessel wall (extrinsic pathway) or by activation of high-molecular-weight kininogen by a number of stimuli. Both pathways lead to the generation of thrombin, an enzyme that turns the soluble blood protein fibrinogen into fibrin, which aggregates into proteofibrils. Another thrombin-generated enxyme, factor XIII, then crosslinks the fibrin ptoteofibrils at the D fragment site, leading to the formation of an insoluble gel which serves as a scaffold for blood clot formation. The circulating enzyme plasmin, the main enzyme of fibrinolysis, cleaves the fibrin gel in a number of places. The resultant fragments, “high molecular weight polymers”, are digested several times more by plasmin to lead to intermediate and then to small polymers (fibrin degradation product or FDPs). The crosslink between two D fragments remains intact, however, and these are exposed on the surface when the fibrin fragments are sufficiently digested. The typical D-dimer containing fragment cotains two D domains and one E domain of the fibrinogen molecule. D-dimer are not normally present in human blood plasma, except when the coagulation system has been activated, for instance because of the presence of thrombosis or disseminated intravascular coagulation. The D-dimer assay depends in the binding of a monoclonal antibody to a particular epitope on the D-dimer fragment. Several detection kits are commerically available; all of them rely on a different monoclonal antibody against D-dimer. Of some of these it is known to which area on the D-dimer the antibody binds. The binding of the antibody is then measured quantitatively by one of various laboratory methods.

Indications

D-dimer testing is of clinical use when there is a suspicion of deep venous thrombosis (DVT) or pulmonary embolism (PE). In patients suspected of disseminated intravascular coagulation (DIC), D-dimers may aid in the diagnosis.

For DVT and PE, there are various scoring systems that are used to determine the a priori clinical probability of these diseases; the best-known were introduced by Wells et al. (2003).

• For a very high score, or pretest probability, a D-dimer will make little difference and anticoagulant therapy will be initiated regardless of test results, and additional testing for DVT or pulmonary embolism may be performed.
• For a moderate or low score, or pretest probability:
  • A negative D-dimer test will virtually rule out thromboembolism: the degree to which the D-dimer reduces the probability of thrombotic disease is dependent on the test properties of the specific test used in your clinical setting: most available D-dimer tests with a negative result will reduce the probability of thromboembolic disease to less than 1% if the pretest probability is less than 15-20%
  • If the D-dimer reads high, then further testing (ultrasound of the leg veins or lung scintigraphy or CT scanning) is required to confirm the presence of thrombus. Anticoagulant therapy may be started at this point or withheld until further tests confirm the diagnosis, depending on the clinical situation.

In some hospitals, they are measured by laboratories after a form is completed showing the probability score and only if the probability score is low or intermediate. This would reduce the need for unnecessary tests in those who are high-probability.

Test properties

Various kits have a 93-95% sensitivity and about 50% specificity in the diagnosis of thrombotic disease.

• False positive readings can be due to various causes: liver disease, high rheumatoid factor, inflammation, malignancy, trauma, pregnancy, recent surgery as well as advanced age
• False negative readings can occur if the sample is taken either too early after thrombus formation or if testing is delayed for several days. Additionally, the presence of anti-coagulation can render the test negative because it prevents thrombus extension.
• Likelihood ratios are derived from sensitivity and specificity to adjust pretest probability.

History

D-dimer was originally described in the 1970s, and found its diagnostic application in the 1990s.