Osmotic gradient ektacytometry is part of the laboratory diagnosis process of hereditary spherocytosis (HS) and other red blood cell (RBC) membrane disorders. We here present the experience of two independent institutions with a next-generation ektacytometer, the LoRRca MaxSis analyzer, in HS diagnostic settings. Inter- and intra-assay variability and sample stability were analyzed. Samples from patients with HS (n=40), probable HS (n=21), auto-immune hemolytic anemia (n=7), and other pathologies (n=37) were studied. Daily controls were run in parallel with patient samples. Results were expressed as percent of change compared to mean of controls. Analytical performances showed an inter-assay variability between 0.2% and 3%. Samples were stable for 48-72 h depending of temperature storage and anticoagulant used. The following diagnostic cut-offs were established for HS: an increase of more than 21.5% for the osmolality point at the minimal elongation index (O min), a decrease of more than 8.5% for the maximal elongation index (EI max), and a decreased area under the curve (AUC) of more than 18.5% compared to the mean of controls. As the previous instrument, the next-generation ektacytometer is an efficient tool for laboratory diagnosis of HS. Sample stability and standardized reporting of results allow inter-laboratory exchange and comparison. The most useful parameters for HS diagnosis were AUC, EI max, and O min; unfortunately, this method does not differentiate between HS and auto-immune hemolytic anemia (AIHA), but it distinguishes HS from other hereditary membrane pathologies. It can thus be considered as an intermediate step between screening and diagnostic tests.