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Abstract
A flow cytometry-based phagocytosis assay was developed and utilized to measure the LE cell phenomenon at the single cell level in vitro. Since the lupus erythematosus (LE) cell phenomenon is a special form of necro-phagocytosis in the presence of anti-dsDNA antibodies, dead substrate cells or chicken erythrocytes nuclei (CEN) served as targets that were labeled with propidiumiodide (PI). Phagocytes (PMN) were stained by anti-CD45 mAb FITC. After co-incubation phagocytosis was measured by flow cytometry. Flow cytometric analysis enabled the discrimination between PI+/CD45- targets, PI-/CD45+ phagocytes, and PI+/CD45+ phagocytes with engulfed targets.
Maintaining the samples on ice significantly reduced the phagocytic uptake as compared to samples co-cultivated at 37°C (p<0.0002). The phagocytic up-take was lowest after substrate pre-treatment in normal serum as compared to samples with either no serum exposure or pre-treatment in LE-serum with anti-dsDNA antibodies (p<0.05).
Taken together, these data suggest the phagocytosis-based flow cytometry assay is suitable for analyzing the LE cell phenomenon. This method provides an interesting, simple and rapid new tool, and will possibly alleviate further studies on the LE cell phenomenon with modified cell models and/or conditions.
