Isolation and some biochemical characteristics of nuclei from AH-66 hepatoma cells

Abstract

Cetylpyridinium chloride uniquely facilitated the isolation of nuclei from AH-66 hepatoma ascites cells in an isotonic medium without homogenization because of its strong solubilization of their plasma membranes, which were resistant to mechanical shearing with the commonly used nonionic detergents such as Triton X-100, Nonidet P-40, and Tween 80. Virtually all the nuclei in a population of AH-66 cells (10(6)/ml) can be isolated with 0.2% cetylpyridinium chloride. The isolated nuclei were free of adherent cytoplasm, maintained satisfactory morphology, and had high activity of nicotinamide adenine dinucleotide pyrophosphorylase. Two-dimensional polyacrylamide gel electrophoresis of the acid-soluble nuclear proteins of the AH-66 hepatoma nuclei isolated by the cetylpyridinium chloride procedure as well as by the citric acid procedure revealed that Spots Ac and C16-C18 were significantly intense in the gel pattern. Unexpectedly, Spot A10 was absent from the gel pattern of AH-66 hepatoma nuclei.