Impact of glycolipid particles in fetal bovine serum on the flow cytometric detection of cell-surface SSEA-4.

We recently established a flow cytometer assay that can detect the interaction between antibodies and glycolipid antigens present on lipid vesicles such as liposomes and extracellular vesicles. Using this system, we detected particles reactive with an anti-stage-specific embryonic antigen-4 (SSEA-4) antibody (MC813-70) in fetal bovine serum (FBS). SSEA-4 is a carbohydrate antigen also known as sialyl-Gb5 glycosphingolipid and is widely used as a marker of human stem cells. We detected an SSEA-4-reactive signal in precipitated particles obtained by centrifugation of FBS by 10,000g (10KgP). However, no SSEA-4-reactive signal was detected in 10KgP prepared from calf serum, suggesting that the detected signal is associated with fetal-derived serum components. Furthermore, we found that an SSEA-4-reactive signal was detected on HeLa cells, which are not known to express SSEA-4, when cultured in a medium containing FBS. By contrast, no SSEA-4-reactive signal was detected on HeLa cells cultured in serum-free medium. These findings indicate that exposure to FBS-containing medium resulted in an apparent SSEA-4-associated surface signal on HeLa cells.