The centrosome of higher eukaryotic cells is the main microtubule-organising centre. To understand the molecular mechanisms underlying this organelle's biogenesis and important functions in several cellular processes, such as microtubule nucleation, cell division and stress response, it was critical to develop methods for isolating biochemically meaningful quantities of centrosomes. Centrosomes have been isolated from a variety of organisms and based on these preparations, numerous aspects of this intriguing organelle's morphological, functional and biochemical properties have been uncovered. Better isolation procedures along with the development of new technologies, like RNAi (ribonucleic acid interference) and the increasing accuracy of mass spectrometry and electron microscopy techniques, have profoundly improved our knowledge of the centrosome, leading to a better understanding of its implications in various cellular processes and in diseases.

Key concepts

• Cultured animal cells, animal tissue, Drosophila embryos, green algae, budding yeast, fungi and surf clams are suitable starting materials for centrosome isolation.
  
• Centrosome isolation from animal cells requires cell lysis in low ionic strength buffers.
  
• Enrichment of centrosomes is achieved by density centrifugation in sucrose step gradients.
  
• Immunoaffinity purification has been employed to further enrich centrosomes from Drosophila embryos.
  
• Isolated centrosomes are competent to nucleate microtubules.
  
• The analysis of isolated centrosomes has provided a wealth of information on their biochemical composition and novel structural and functional features.
  
• Many centrosome components have been identified by applying mass spectrometry and antibody approaches.
  
• RNAi has been used to study centrosome functions in cellular signalling pathways and the cell cycle and to identify novel components.
  
• Electron tomography will be an informative tool to study the supramolecular structures of isolated centrosomes.