sHLA has been described in human serum and other body fluids. In this study sHLA shed by cultivated human cells and their biochemical nature in solution were studied. EBV-transformed human B-lymphoblastoid cell lines (n = 4), permanent human lymphoblastoid tumor cell lines (n = 4), and PBLs from three donors were cultivated in vitro and sHLA measured in the supernatants. The Daudi cell line was used as a negative control in all experiments. Maximum expression of sHLA was measured after 8 hours, after which the concentrations gradually declined. The allospecificities A2 and B7 were also detectable in the ELISA. sHLA in the supernatants was further characterized by 1D-IEF. All bands representing the allotypes were detected, showing that cell supernatants can be used as antigen sources for biochemical tissue typing. These data show that sHLA expression is a characteristic of viable cells.