PURPOSE: To establish new in vitro model systems that better reflect in vivo condition, multicellular tumor spheroids(MTS) and raft culture were developed using cell lines of squamous cell carcinoma(SCCHN) of the head and neck. In these 3-dimensional systems, the expression of cell surface molecules which are important for modulation of physiology of tumor cells were studied with or without the treatment of interferon(IFN)-gamma. MATERIALS AND METHODS: Four SCCHN cell lines were used for MTS and raft culture. The effects of interferon-gamma on SCCHN cells were examined by immunohistochemistry. RESULTS: All cell lines formed MTS, but only Tu-138 showed a good stratification at the air-liquid interface in the raft culture system. Immunohistochemical studies of MTS using monoclonal antibodies revealed a strong staining for MHC class I, no staining for MHC-DR, a weak patch expression of ICAM-1 and a central strong staining for integrin a 6. Staining patterns were similar for the raft cultures except integrin a 6(intense full-thickness positivity). In both systems, IFN-gamma enhanced the expression of MHC-DR and ICAM-1. No significant change was found in the expression of MHC class I and integrin a 6. CONCLUSIONS: MTS and raft culture system were established successfully from the SCCHN cell lines. IFN-gamma can modulate the surface molecules of tumor cells in the 3-dimensional culture systems.