Non-selective isolation of human somatic cell hybrids by unit-gravity sedimentation

DATA from somatic cell hybridisation have made important contributions to the understanding of genetic control of pheno-typic expression, the nature of malignancy in tumours, gene mapping and the basic genetic lesions causing metabolic diseases1–5. Broader application has been limited by the absence of biochemical markers in most cell strains and lines, making it difficult to eliminate parental cells and isolate hybrids. Recent non-selective methods of isolating hybrid cells require either expensive and elaborate equipment6 or examination of large numbers of clones with a low yield of hybrids7. We now present a method for isolating tetraploid clones from fused human fibroblasts non-selectively and in high yield. Our technique is applicable to tetraploid cells formed from any pair of diploid cells as long as the synkaryons so formed can proliferate. The method depends on the separation of fused cells from the unfused parental cells because of their greater size. A mixture of cells settles out according to size in a shallow bovine serum albumin (BSA) gradient at unit gravity, yielding fractions enriched in double-nucleated cells from which tetraploid clones can be isolated in high yield.