Enrichment of human heterokaryons by Ficoll gradient for complementation analysis of iduronate sulfatase deficiency.
Ficoll gradients have been used to enrich for heterokaryons in cultures of human skin fibroblasts following polyethylene glycol (PEG) induced fusion. These gradients provide a simple and consistent method for obtaining populations of multinucleated cells, at least twofold greater than those resulting from fusion alone. Formation of glucose-6-phosphate dehydrogenase (G6PD) heteropolymers has been used as a functional assay for the presence of heterokaryons. Analysis of cell populations enriched for multinucleated cells has revealed complementation leading to iduronate sulfatase activity in heterokaryons derived from iduronate sulfatase-deficient fibroblasts expressing the Hunter and multiple sulfatase-deficiency mutations.
D002499:Centrifugation, Density Gradient / Q000379:methods*; D005362:Ficoll; D005816:Genetic Complementation Test; D005954:Glucosephosphate Dehydrogenase / Q000235:genetics; D006801:Humans; D006822:Hybrid Cells / Q000201:enzymology; D007066:Iduronate Sulfatase / Q000235:genetics*; D016532:Mucopolysaccharidosis II / Q000235:genetics; D010641:Phenotype; D012867:Skin / Q000166:cytology / Q000201:enzymology; D013429:Sulfatases / Q000235:genetics*
D005362:Ficoll; D005954:Glucosephosphate Dehydrogenase; D013429:Sulfatases; D007066:Iduronate Sulfatase