The identification of mechanisms maintaining or even rebuilding the endothelial glycocalyx (eGC) are highly desirable. Currently, the evidence about mechanisms for protection and maintenance of the eGC is low. EGC preserving factors are for example albumin or the high-density lipoproteins (HDL) with their bound sphingosine-1 phosphate. An interesting approach was demonstrated by Machin et al. They demonstrated that dietary supplementation of glycocalyx precursors as in Endocalyx Pro™ are able to enhance the eGC barrier function in older mice (7). Pilot data in apparently healthy individuals show a significant up to 10% improvement of eGC dimensions after a short oral Endocalyx intake in suggested dosing. A clinical study is currently testing the effect of the Endocalyx Pro™ in the context of diabetic nephropathy (Clinicaltrials.gov: NCT03889236). Endocalyx Pro™ preserves and refurbishes eGC in vitro after enzymatic digestion. Whether Endocalyx Pro™ preserves eGC in vitro has not been investigated yet. Therefore, we analysed the effect of Endocalyx Pro™ in endothelial cell culture and quantified eGC with the atomic force microscopy (AFM). Our group has successfully established AFM methodology to quantify nanomechanical endothelial cell properties, i. e. stiffness and thickness, of the eGC. AFM is able to reveal the existence of a mature eGC on the luminal endothelial surface of freshly isolated rodent aorta preparations ex vivo as well as on a variety of living endothelial cells in vitro. Preliminary results indicate that Endocalyx Pro™ is able to preserve the eGC in vitro. Interestingly, Endocalyx Pro™ is not only capable of maintaining the eGC, but to refurbish eGC-thickness after enzymatic digestion by heparinase 1. Further experiments are needed to rule out the mechanisms by which Endocalyx Pro™ enhances eGC thickness. Furthermore, the decrease of eGC-height caused by 5% serum of hemodialysis patients was reversed by Endocalyx Pro™ incubation as well.