Beckman Coulter Application (system and method support)
Method for the Purification of Eukaryotic Ribosomes
Allen Furst
Beckman Coulter, Inc.
Ribosomes may be prepared from cell lysates by pelleting at 100,000 x g, and the ribosomal subunits further separated by rate zonal sedimentation on a sucrose gradient. Typical starting materials are reticulocyte lysates, wheat germ extracts, or E. coli. The absence of nuclei in these materials facilitates the initial isolation of polysomes. The method for rabbit reticulocytes given here is based on that of Merrick(1). It provides ribosomes suitable for translational assays and for proteomics studies. To preserve activity, it is best to complete the procedure as rapidly as possible. See also Beckman Coulter Application Bulletin A-1850A ("A Rapid Method for Ribosome Preparation: Part 1-- Using the High-Capacity Fixed Angle MLA Rotor in an Optima™ MAX Tabletop Ultracentrifuge") for isolation of bacterial ribosomes.
References:
- Merrick, W. C. (1979) "Purification of protein synthesis initiation factors from rabbit reticulocytes." Methods in Enzymology 60: 108-123.
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