Tomato farms in Arusha, Morogoro, Dodoma, Iringa, Kilimanjaro and Coast regions of Tanzania were surveyed to assess the incidence of the yellow leaf curl disease, and to collect infected tomato leaf samples for sero-diagnosis. The triple antibody sandwich enzyme linked immunosorbent assay (TAS-ELISA) format was adopted for the detection of disease using commercial polyclonal antiserum and monoclonal antibodies SCRI 17, SCRI 20, SCRI 23 and SCRI 33. ELISA readings were rated on a scale of 0–4. The results of the tests indicated that all the Tomato yellow leaf curl virus (TY-LCV) isolates recorded high reaction values (4) with the polyclonal antibody. However, the Dodoma and Arusha isolates were rated highest in optical density (OD) reading with MAb SCRI 20 and 23. The remaining isolates produced lower OD values. All the isolates rated low (2) when tested with SCRI 33. The differences in reaction to the monoclonal antibodies of TYLCV indicated that variability exists between the coat protein epitopes of TYLCV and Tomato yellow leaf curl Tanzania virus (TYL-CTZV) on one hand, and among the TYLCTZV isolates on the other. Only the isolates from Arusha and Dodoma share a high sequence homology in coat protein with the European and related TYLCV isolates. Furthermore, the reaction with either SCRI 20 or SCRI 23 show that the isolates from Arusha and Dodoma share a high degree of homology, and could belong to one serotype. The other isolates from Morogoro, Coast and Kilimanjaro could form another serotype, while the isolate from Iringa is a different serotype. On the other hand, reaction with SCRI 17 groups the isolates in two serotypes, the Dodoma isolate alone, and another that groups the other five isolates together. It is recommended that other procedures such as DNA-DNA hybridization assays, polymerase chain reaction, restriction fragment length polymorphisms and sequencing can be combined with the use of monoclonal antisera for the detection and prediction or inference of Tomato yellow leaf curl disease (TYLCD) virus relationships at the quasi-species or strain levels in Tanzania