The large T protein coded by the early region of simian virus 40 (SV40) is involved in the induction and maintenance of cell transformation. It is not clear which properties of T are important in causing the transformation, since the protein is multifunctional. To clarify the action of T proteins in transformation, we have prepared several monoclonal antibodies directed against different regions of the T molecule. One of these antibodies, DL 3C4, recognizes an antigenic determinant on T that is also present on a host cell protein of molecular weight 68,000. This cross-reactive 68K protein is located within the nucleus of all mammalian cell types examined and has a characteristic granular distribution as shown by immunofluorescence. The antigenic determinant recognized by DL 3C4 is resistant to denaturation. Studies on Adeno-SV40 hybrid viruses show that the antigenic site on T is coded for by sequences located between 0.44 and 0.29 on the SV40 map.