May 2009

Latest in Haematology Analyzers:Fluorescence Flow Cytometry

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Haematology has evolved into an interesting subject due to the advanced technology. Also, over the past few years; the market penetration of haematology analyzers has increased significantly in laboratories across the country. The latest technology in haematology analyzers is Fluorescence Flow Cytometry (FCM).

The term “cytometry” is defined as a measurement of physicochemical properties of cells and other biological particles. Flow cytometry offers measurements of cells and other particles flowing in thin streams. Generally, it detects optical information from cells or other particles flowing in a thin stream under irradiation of a laser beam. Such optical information sources include scattered light and fluorescence depending on the measurement objective.

Since the late 1990s, cell counter manufacturers have been looking at flow cytometry techniques as a way of increasing the capabilities of cell counters as traditional flow cytometry instrumentation requires the use of highly specific antigen-antibody reactions; often requiring pre-incubation and use of expensive reagents. In addition, gating of cell scattergram is complex which is manually performed by skilled operator, complexities of the reagents used, requirements of manual pipetting and not all applications are automated.

Haematology analyzers with FCM use Florescence dye to stain all sampled cells, which increases specificity and allows extension of clinical applications beyond the realm of traditional cell counting, without the complexity and cost of antigen/antibody reactions.

Recently, ingenious Fluorescence Flow Cytometry (FCM) Technology platform is introduced in mid-size haematology analyzers, which was successfully introduced with top-of-the-end haematology analyzers. These analyzers with FCM offer excellent capabilities of best differentiation of normal cells and pathological cells; and interference is managed better that before and that also without compromising throughput and reliability.

The major benefit of FCM to its users is improved detection of pathological cells like Immature Granulocyte Count, High Fluorescence Lymphocyte Count, Abnormal Lymphocyte Detection etc. The other advantage is improved quality of 5 part differential of WBC with 48 to 72 hours stability after blood collection and that also for High linear ranges (no dilution necessary). It has a stable and long life detector system requiring no calibration.

As depicted in the image of Optical System of haematology analyzers with FCM, it detects each cell from three angles.

  • Forward Scattered Light (Information on Cell volume)
  • Side Scattered Light (Information on Internal Cell structure)
  • Side Fluorescence Light (Information on RNA/DNA content)

Fluorescence Light detects DNA/RNA information of cells along with information about Cell activity such as Duplication activity of the nucleus (High RNA) &  Cytoplasm activity (Protein synthesis etc)

This information is useful for very good differentiation of mature and immature cells. With use of artificial intelligence based complex computer algorithm this differentiation is plotted as easy to interpret scattergram by the haematology analyzers.

Hence, FCM enables Haematology analyzers to differentiate not only mature WBCs, but also mature and immature WBCs. By staining the cells, analytical sensitivity of cell counter is enhanced and signal-to-noise ratio is increased. Reportable ranges are extended and interferences are reduced. FCM enabled Haematology analyzers are now able give more detailed IP messages and suspect messages like Neutropenia, Neutrophilia, Lymphopenia, Lymphocytosis, Monocytosis, Eosinophilia, Basophilia, Leukocytopenia, Leukocytosis, NRBC Present,  Blasts, Immature Granulocytes, Abnormal  Lymphoblasts, lymphocytes, NRBC, RBC Lyse Resistance, Atypical Lymphocytes, Anisocytosis, Microcytosis, Macrocytosis, Hypochromia, Anemia, Erythrocytosis and Reticulocytosis

They are also able to analyze special parameters like Fluorescence Platelet (PLT-FL), immature Granulocytes (IG), Nucleated Red Blood Cell (NRBC), Hematopoietic Progenitor Cells (HPC), Reticulocyte, Immature Reticulocyte Fraction (IRF), Reticulocyte Haemoglobin (RET-He),   Immature Platelet Fraction (IPF), etc. Analysis of these parameters give insight into bone marrow erythropoietic activity, timing for apheresis in peripheral blood stem cell transplantation, clinical information of peripheral platelet destruction and marrow failure in thrombocytopenic patients

(Article contributed by Transasia Bio-Medicals Ltd.

E-mail : transasia@transasia.co.in
Website : www.transasia.co.in)

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