Hematology Lessons Best Lab: Flow Cytometry-based Hematology Analyzers

Hematology Analyzers

Background Information of Hematological Analyzers

Hematology analyzers are computerized, highly specialized machines that count the number of different types of red and white blood cells, blood platelets, haemoglobin, and haematocrit levels in a blood sample. They include programmable automated alarm systems for indicating results outside the reference range. Each hematological anaylzer has their own testing principles to derive with their cell counts and parameters. One of the most common principle is flow cytometry. 

Flow Cytometry


Background Information of Flow Cytometry

The term ‘cytometry’ is defined as a measurement of the physico-chemical properties of cells. ‘Flow cytometry’ is the measurement of cells and other particles in a flow.Traditional flow cytometry has always been considered the best method to distinguish and differentiate the various cell populations. However, the costly need to use fluorescence-labelled antigen-antibody reagents and the cumbersome tedious preparation procedures is a major disadvantage. In order to meet the clinical demands, some companies (Eg Sysmex) have professionally refined basic techniques of flow cytometry and uses highly specific polymethine dyes to adapt this sophisticated technology to fit the high volume and automation demands of clinical laboratories.

Test Principle of Flow Cytometry

Once a cell/object comes into the path of a ray of light (Laser), the light changes its orientation. This phenomenon is called light scatter and occurs under all angles between 0 and 360°. 

Detection of this scattered light provides information on the size and the quality of the object. Especially light scattered in forward direction (‘forward scattered light’) or, respectively, low-angle scattered light allows statements on the size of an object. Low-angle scattered light is decisively influenced by the object’s form and refraction index, aside from its size.

On the other hand, light scattered from the side (‘side scattered light’) or wide-angle forward scattered light provides information on the internal granularity content of a cell/object. That way, blood cells can be analysed with regard to their size and their intracellular, structural complexity (form, size and density of the nucleus, cytoplasmic granularity). Wide-angle forward scattered light is decisively influenced by the presence or absence of granules.

A particularly sensitive detector is required for the detection of side scattered light – as
a rule a photo multiplier – since, compared with forward scattered light, the signal intensity is weaker by some powers of ten. A fast photo diode can be used for the forward scattered light. 

Under specific prerequisites, the exact size of particles can be determined by means of the light scatter method. Furthermore successful will be the excellent separation of different particle populations based on additional properties such as the surface of the cell.

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