Old and new insights into the diagnosis of hereditary spherocytosis

Abstract

Hereditary spherocytosis (HS) belongs to the group of congenital hemolytic anemias resulting from plasma membrane protein deficiency. When diagnosed too late, HS bares the risk of long-term complications including gall stones and severe anemia. Here, there are discussed advances in HS screening and diagnostics, with a particular focus on methodologies, most of which are available in clinical laboratories worldwide.

Keywords: Complete blood count (CBC); ektacytometry; eosin-5’-maleimide; hereditary spherocytosis (HS).

Figure 1

Flow cytometric osmotic fragility test (FCOF). Ratio between cells count in first (P1) and mean of two last gates [(P9+P10)/2]corrected by dilution factor indicates residual red blood cells count, which, when low, is indicative for HS. (A) Gating of red blood cells in forward scatter vs. side scatter logarithmic scale cytogram; (B) cytogram of forward scatter vs. time divided into 30 seconds gates, P1—shows cells before water addition into red blood cells suspension in saline, and gates P2–P10 show cells after water addition; (C) cells counts in appropriate gates.

Figure 2

Eosin-5’-maleimide test. (A) Red blood cells are gated in gate “RBCs”, gate “E” contains doublets of erythrocytes. Primary gating is based on forward scatter vs. side scatter logarithmic scale; (B) EMA-bound cells’ fluorescence is measured in the first channel of fluorescence at 488 nm excitation wavelength and presented as mean fluorescence intensity; (C) the overlay histogram of fluorescence of EMA-stained RBCs from the HS subject (first, lower peak) and a healthy control (second peak), respectively. Peaks differ in mean fluorescence intensity.