Three new monoclonal antibodies (MAbs) (MOv16, MOv18 chemical characterization of 3 MAbs obtained using this novel and MOv 19) were raised against human ovarian carcinoma. approach, 2 of which show tumor-restricted reactivities.To obtain more specific reagents than those produced so far, we adopted the following experimental approach which consisted of: (I) the selection of a poorly differentiated ovarian carcinoma which was unreactive with all the MAb previously cell lines selected in our laboratory; and (2) the application of a particular immunization protocol. The reactivity of the selected Human Cell lines including ovarian carcinoma SW-626, co-MAbs was studied by solid-phase RIA on live and fixed cells lon Carcinoma HT-29, mammary carcinomas MCF-7 and SKfrom tumor cell lines and by immunofluorescence on frozen Br-3, pancreatic carcinoma CAPAN-1, melanoma HT-144 and sections from surgical specimens. The MAb MOvI6 reacted Sarcoma SW-684 were all provided by Dr. J. Fogh (Memorid with 60% of ovarian carcinomas as well as with a high Per-Sloan-Kettering Cancer Center, New York, NY). The ovarian centage of other carcinomas and with some normal tissues. In carcinoma cell lines ~~-0 ~4 and ovca432 and the lung carcontrast, MOv18 and MOv19 appeared to have restricted specon other carcinomas was only observed in a few cases and no K.L. Lloyd (MSKCC), Dr. R. KnaPp (Dana Farber Institute, reactivity was found on non-epithelial tumors or normal tis-Boston, MA) and J.D. Minna (Naval Institute, Bethesda, sues. lmmunoprecipitation experiments indicated that MOv16 MD). The BALB/c myeloma cell line P3-NS1 (NS1) was used recognizes a 48-50-kDA protein, whereas MOv18 and MOvl9 for production of the hybridomas. The cell lines were all both identify a 38-40 kDA glycoprotein band. Cross-competi-maintained in RPMI-1640 medium (Microbiological Assocition experiments, together with a double-determinant im-ates, Walkersville, MD) supplemented with 10% fetal calf munoradiometric assay which uses MOv18 as catcher and Serum (FCS), penicillin (100 pg/ml) and streptomycin MOv 19 as tracer, suggested that they recognize different epitopes carried by the same molecule. The affinity constants of pg'ml)' MOv18 and MOvl9 were estimated to be in the range of 10' ~~-1 cell suspensions -lo9 M-'. Taken together, the properties of these antibodies, their restricted ovarian tumor specificities and relative and peripheral blood lymphocytes were high affinity constants, suggest that they could represent obtained respectively from non-carcinoma patients and healthy promising tools for in vivo applications.