The addition of insulin to HAT medium (HIAT) enhances hybridoma formation.

To assess the effects of insulin on the formation of hybridomas, sheep red blood cell (SRBC) immunized spleen cells from BALB/c mice were fused with P3U1 mouse myeloma cells. After fusion, cells were grown for 2 weeks in HAT medium containing insulin (HIAT) (doses ranging between 10(-1) to 10(-9) units/ml) or HAT medium alone. The number of hybridoma colonies was found to be significantly increased in the presence of HIAT medium compared to HAT alone. In addition, the average size of the hybridoma clones was at least doubled and the cumulative colony size index per plate increased several folds. A significant rise in the number of wells containing clones secreting anti-SRBC monoclonal antibodies was again shown in the presence of HIAT compared to HAT medium alone. The maximal effect of insulin on the above biological parameters ranged between 10(-1) and 10(-4) units/ml. It is concluded that the addition of insulin to HAT medium (HIAT) enhances hybridoma formation and thus, its more frequent use may considerably expediate ongoing research efforts on the production of monoclonal antibodies.