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. 2020 May;98(3):238-249.
doi: 10.1002/cyto.b.21857. Epub 2019 Nov 21.

Applications of imaging flow cytometry in the diagnostic assessment of red cell membrane disorders

Tejashree Anil More  1 , Bhavik Dalal  1 , Rati Devendra  1 , Prashant Warang  1 , Aruna Shankarkumar  1 , Prabhakar Kedar  1
Affiliations
Free article

Applications of imaging flow cytometry in the diagnostic assessment of red cell membrane disorders

Tejashree Anil More et al. Cytometry B Clin Cytom. 2020 May.
Free article

Abstract

Background: Red cell membranopathies refers to phenotypically and morphologically heterogeneous disorders. High throughput imaging flow cytometry (IFC) combines the speed, sensitivity, and phenotyping abilities of flow cytometry with the detailed imagery and functional insights of microscopy to produce high content image analysis with quantitative analysis. We have evaluated the applications of IFC to examine both the morphology as well as fluorescence signal intensity in red cell membranopathies.

Methods: Fluorescence intensity of eosin-5-maleimide (EMA) labeled red cells was measured for diagnosis of RBC membrane protein defect on Amnis ImageStreamX followed by Image analysis on IDEAS software to study features such as circularity and shape ratio.

Results: The hereditary spherocytosis (HS) group showed significantly decreased MFI (52,800 ± 9,100) than normal controls (81,100 ± 4,700) (p < .05) whereas non-HS showed 78,300 ± 9,900. The shape ratio of hereditary elliptocytosis (HE) was significantly higher (43.8%) than normal controls (14.6%). The circularity score is higher in HS (64.15%) than the normal controls (44.3%) whereas the circularity score was very less in HE (10%) due to the presence of elliptocytes.

Conclusions: The advantages of the IFC over standard flow cytometry is its ability to provide high-content image analysis and measurement of parameters such as circularity and shape ratio allow discriminating red cell membranopathies (HS and HE) due to variations in shape and size. It could be a single, effective, and rapid IFC test for detection and differentiation of red cell membrane disorders in hematology laboratories where an IFC is available.

Keywords: hereditary spherocytosis; imaging flow cytometry; red blood cell.

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References

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