Flow Cytometry its uses -Are we aware??

What is Flow Cytometry??  Common blood tests like say, complete haemogram , types of cells myeloid series, Erythroid series, RDW are measured by instruments based on either impedance technology or impedance technology ,This includes RDW as well

Flow Cytometry:  a costly machine  is designed for  recognition and final diag of  Solid   tumors ,lymphoma, leukemia,  some genetic mutations. Genetic mutations. Its indications in our discipline is limited,  Flow cytometry measures optical and fluorescence characteristics of single cells (or any other particle, including nuclei, microorganisms, chromosome preparations, and latex beads).

 

 

, Physical properties, such as size (represented by forward angle light scatter) and internal complexity (represented by right-angle scatter) can resolve certain cell populations. Fluorescent dyes may bind or intercalate with different cellular components such as DNA or RNA. Additionally, antibodies conjugated to fluorescent dyes can bind specific proteins on cell membranes or inside cells. When labeled cells are passed by a light source, the fluorescent molecules are excited to a higher energy state. Upon returning to their resting states, the fluorochromes emit light energy at higher wavelengths. The use of multiple fluorochromes, each with similar excitation wavelengths and different emission wavelengths (or “colors”), allows several cell properties to be measured simultaneously. Commonly used dyes include propidium iodide, phycoerythrin, and fluorescein, although many other dyes are available. Tandem dyes with internal fluorescence resonance energy transfer can create even longer wavelengths and more colors. Table 1⇓ lists clinical applications and cellular characteristics that are commonly measured.

 

The use of flow cytometry in the clinical laboratory has grown substantially in the past decade. This is attributable in part to the development of smaller, user-friendly, less-expensive instruments and a continuous increase in the number of clinical applications. Flow cytometry measures multiple characteristics of individual particles flowing in single file in a stream of fluid. Light scattering at different angles can distinguish differences in size and internal complexity, whereas light emitted from fluorescently labeled antibodies can identify a wide array of cell surface and cytoplasmic antigens. This approach makes flow cytometry a powerful tool for detailed analysis of complex populations in a short period of time. This report reviews the general principles in flow cytometry and selected applications of flow cytometry in the clinical hematology laboratory.

Flow cytometry provides rapid analysis of multiple characteristics of single cells. The information obtained is both qualitative and quantitative. Whereas in the past flow cytometers were found only in larger academic centers, advances in technology now make it possible for community hospitals to use this methodology. Contemporary flow cytometers are much smaller, less expensive, more user-friendly, and well suited for high-volume operation. Flow cytometry is used for immunophenotyping of a variety of specimens, including whole blood, bone marrow, serous cavity fluids, cerebrospinal fluid, urine, and solid tissues. Characteristics that can be measured include cell size, cytoplasmic complexity, DNA or RNA content, and a wide range of membrane-bound and intracellular proteins. mmunophenotyping

The most common application performed on the cytometer is immunophenotyping. This technique identifies and quantifies populations of cells in a heterogeneous sample - usually blood, bone marrow or lymph. These cell subsets are measured by labeling population-specific proteins with a fluorescent tag on the cell surface. In clinical labs, immunophenotyping is useful in diagnosing hematological malignancies such as lymphomas and leukemia.

Cell Sorting

The cell sorter is a specialized flow cytometer with the ability to physically isolate cells of interest into separate collection tubes. The cytometer interrogates and characterizes each cell as it passes through the laser. The sorter then uses sophisticated electronics and fluidics to identify and "kick" the cells of interest out of the fluidic stream into a test tube.

Cell Cycle Analysis

Flow cytometry can analyze replication states using fluorescent dyes to measure the four distinct phases of the cell cycle. Along with determining cell cycle replication states, the assay can measure cell aneuploidy associated with chromosomal abnormalities.

Apoptosis

Apoptosis, or programmed cell death, is a normal part of the life cycle of eukaryotic cells. Cells die for a variety of reasons: through necrosis, brought on by external physical and chemical changes to the cell or through apoptosis, a process in which cells initiate a "suicide" program through internally controlled factors. These two distinct types of cell death, apoptosis and necrosis, can be distinguished by flow cytometry on the basis of differences in morphological, biochemical and molecular changes occurring in the dying cells.

Cell Proliferation Assays

Cell proliferation assays are widely used in cell biology to measure cellular metabolic activity in response to stimuli such as growth factors, cytokines and other media components. The flow cytometer can measure proliferation by labeling resting cells with a cell membrane fluorescent dye, carboxyfluorescein succinimidyl ester (CFSE). When the cells are activated, they begin to proliferate and undergo mitosis. As the cells divide, half of the original dye is passed on to each daughter cell. By measuring the reduction of the fluorescence signal, researchers can calculate cellular activation and proliferation.