Journal of Immunological Methods, 104 (1987) 1-6 1 Elsevier JIM 04501 Rapid dissociation of adherent human tumor cells by ultrasound Hans D. Menssen, Meenhard Herlyn, Ulrich Rodeck and Hilary Koprowski The Wistar Institute of Anatomy and Biology, 36th Street at Spruce, Philadelphia, PA 19104, U.S.A. (Received4 May 1987, accepted 22 May 1987) Cultured human melanoma and gastrointestinal carcinoma cells were detached from substrate and further dissociated by placing the culture vessel into a water-filled ultrasonic cleaner (43 kHz) and sonicating it for 10-50 s. Plating efficiency and long-term growth of three melanoma cell lines were similar after ultrasound or trypsin detachment. Binding of monoclonal antibodies that define normal and tumor-associated antigens on melanoma and colorectal carcinoma cells was not affected by ultrasound in 21 out of 23 cases. The 40 kDa colorectal carcinoma-associated antigen defined by monoclonal antibody CO 17-1A was more highly expressed after ultrasonication than trypsinization. The antigen defined by antibody CO 44.1 on these cells was more sensitive to sonication. This method represents a rapid, effective and gentle alternative to trypsin detachment of cultured cells, especially when repeated cell washing or centrifugation steps are required. Key words: Ultrasound detachment in vitro; Melanoma; Colorectalcarcinoma; Antigen expression Introduction Recent advances in tissue culture techniques have allowed the culture of many different normal and malignant human ceils in chemically defined media. The absence of undefined components that are present in fetal bovine serum (FBS) facilitates immunological studies such as antigen purification or modulation of antigen expression. The growth of cells in serum-free medium is laborious since routine passaging of cells by tryp- sinization requires additional centrifugation steps. Furthermore, cells maintained in serum-free medium or with low serum concentrations may also be more susceptible to trypsin digestion than cells maintained in medium with high FBS con- centrations. Trypsin sensitivity of surface antigens has been reported for several cultured tissues (Molinari and Platt, 1975; Friedlander and Fisch- man, 1977). Here we report a fast, simple and inexpensive technique to efficiently detach adherent human tumor cells from substrate by short exposure to ultrasound. Correspondence to: M. Herlyn, The Wistar Institute of Anatomy and Biology,36th Street at Spruce, Philadelphia,PA 19104, U.S.A. Abbreviations: CEA, carcinoembryonic antigen; EGF, epi- dermal growth factor; EDTA, ethylenedinitrilotetraacetic acid; FBS, fetal bovine serum; FITC, fluorescein isothiocyanate; HBSS, Hanks' balanced salt solutions; MAb, monoclonalanti- body. Materials and methods Cell lines and culture conditions WM 9, WM 164, and WM 852 metastatic melanoma lines have been characterized (Herlyn et al., 1980, 1985b). SW 707 colorectal carcinoma cell line was obtained from A. Leibovitz, Temple, 0022-1759/87/$03.50 © 1987 ElsevierScience Publishers B.V. (Biomedical Division)