SummaryThe spleen is prone to both physical damage and functional impairment, which can be difficult to detect before catastrophic complications occur. Currently available tests of splenic function are laborious, user‐dependent and unreliable, so there is an unmet need for a reliable test offered routinely in diagnostic laboratories. In this study, we have assessed a simple flow cytometry‐based method measuring high mannose glycans (HMGs) on erythrocytes, which has previously been proposed as a potential test of splenic function. We developed the test as a diagnostic assay using blood from a range of control and potentially hyposplenic samples, including people with sickle cell disease. HMG expression correlated well with manual pit counting, an established method for assessing splenic function (r = 0.6). A threshold of >36% difference compared to the mean of control samples was used to define hyposplenism. At this threshold, the test is 93% sensitive and 100% specific for detecting splenic dysfunction. The test was highly reproducible and stable in blood samples of up to 4 days old. This test is non‐invasive with quantitative data output and requires significantly less operator time than other available techniques, making it a robust new clinical assay for determining splenic function.