Iodixanol (Optiprep), an improved density gradient medium for the iso-osmotic isolation of rat liver peroxisomes

The suitability of Iodixanol (5,5'-C(2-hydroxy-1,3-propanediyl)-bis(acetylamino)] bis-[N,N'-bis(2,3-dihydroxypreopyl-2,4,6-triiodo-1,3-benzenecarboxamide)]}, a nonionic iodinated compound with a molecular weight of 1550, for the isolation of peroxisomes from rat liver was investigated Centrifugation of light mitochondrial fractions in 20 to 40% (w/v) Iodixanol gradients, made isoosmotic by the addition of sucrose, resulted in an excellent separation of peroxisomes hom the remaining organelles, which were not able to enter the gradient. Peroxisomes banded around 30% (w/v) Iodixanol (d similar to 1.175) and, as revealed by marker enzyme analysis, were enriched 35- to 40-fold Morphological examination of the peroxisomal fractions confirmed the near absence of other organelles and revealed structurally well-preserved peroxisomes. Free cores, also present in the starting fractions, migrated to higher densities and were trapped on a cushion No interference of Iodixanol with marker enzyme determinations was observed, except for the UV-metric determination of mate oxidase and for the analysis of protein. (C) 1996 Academic Press, Inc