Recovery of heterokaryons at high cell densities following isolation using irreversible biochemical inhibitors.

The selection of heterokaryons using irreversible biochemical inhibitors provides an effective method for isolating fusion products between any types of cultured cells. One weakness of the technique is that the surviving heterokaryons can only be obtained at low culture densities, making their analysis cumbersome. This report describes the use of Ficoll-sodium diatrizoate gradients to partially separate viable from nonviable cells and characterizes the effects of initial plating density and medium volume on heterokaryon survival. The combined result of optimizing these parameters increases cell rescue and permits heterokaryons to attach at densities of 60,000 cells/cm 2 instead of 600 cells/cm 2. These modifications substantially improve the ease of using irreversible biochemical inhibitors to isolate purified populations of heterokaryons.