Flow cytometry

Flow cytometry in Primary Immunodeficiency and Leukemia

Background

Flow cytometry tests enable scientists to establish what particular type of leukaemia a patient has by checking their blood cells for different sets of surface molecules that typify different leukemias

Flow cytometry in the cellular laboratory is largely divided into two areas; primary immunodeficiency and leukaemia diagnosis. For the purposes of diagnosing immunodeficiency, blood cells are checked for the presence of intact surface receptors that mediate effective immune responses. Flow cytometry allows scientists to label specific surface receptors on different populations of blood cells so that this functionality can be investigated. In leukemia a mutation in genetic expression leads to the uncontrolled proliferation of a single lineage of immune system cells. Flow cytometry enables scientists to establish what particular type of leukaemia a patient has by checking their blood cells for different sets of surface molecules that typify different leukemias. It is important to characterise different leukemias because the type can have a significant affect on patient prognosis and treatment.

Science

Flow cytometry results are represented graphically; cells are recorded as having a specific signature based on size, granularity, and abundance of surface markers

Individual populations of immune system cells with specific roles and at different stages of maturation have different groups of molecules on their surface. This characteristic is useful for quantifying the different populations as we can identify them based on these specific markers. Commercially available antibodies that have specificity for single unique surface molecules are mixed with the patient’s blood so that they can bind to their targets. These antibody labels are joined to dyes (fluorochromes) that emit light at specific wavelengths when excited by light at a lower wavelength, several different colours attached to antibodies with different specificities can be used in the same sample. Cells tagged with the different antibodies are rapidly passed in single file through a laser beam that excites the dye molecules. The light that is emitted at different wavelengths is detected through a series of finely aligned detectors. In addition the cell itself casts a unique shadow as it passes through the laser beam. This is recorded and these measurements are extrapolated to determine the size and granularity of the cell. The results are represented graphically and by using this data every cell is recorded as having a specific signature based on size, granularity, and abundance of surface markers.

In some primary immunodeficiencies, certain populations of cells are absent or may have crucial receptor molecules missing on their surfaces. Flow cytometry can be used to check if these populations are present and if they have their full complement of receptor and activation molecules. Leukemia is typified by an abundance of single type of cell at one specific stage of development undergoing rapid proliferation this means that it will be the major type of cell in the sample and in turn results in a reduction of all other cells. Using flow cytometry, this population can be determined as a homogeneous group of cells with a discrete set of surface marker characteristics specific for the type of leukemia.

Clinical

A multi-disciplinary approach,using several departments to confirm diagnosis improves the chances of providing a correct diagnosis so that the patient can receive the appropriate treatment

Leukaemia is a disease of the blood or bone marrow resulting from the cancerous proliferation of cells of the immune system, caused by a mutation in a single stem cell. Patients can present with a variety of symptoms, some of the more common include bone pain, excessive bruising, anemia and uncontrolled infections. Chronic leukaemias will cause death, if untreated, in months or years however acute leukaemias will cause death, if untreated, in weeks or months. A multi-disciplinary approach, utilising flow cytometry in conjunction with clinical history, morphology, full blood count and cytogenetics to diagnose specific types of leukemia is routinely employed. Using several departments to confirm diagnosis is important because it improves the chances of providing a correct diagnosis so that the patient can receive the appropriate treatment as soon as possible.