Centrifuge Theory and Safety

1. Centrifuge Theory and Safety

2. Centrifuge • A mechanical device that uses the principle of centrifugal force to separate materials of different densities. • In biotechnology research, these materials are usually cells, organelles, or large molecules, such as DNA. • Centrifugal force- An apparent force that acts outward on a body moving around a center, arising from the body's inertia (whereas centripetal force is inward)

3. Centrifuge at NASA Ames Research Center. Featured in movie “Space Cowboys,” 2000

4. Urania Observatory, The Netherlands

5. Benchtop Centrifuge • Also known as multipurpose centrifuge. Not always found on a benchtop. • Uses: Pellet cells and bacteria • Fixed angle rotor: 17,000 g/14,000 rpm • Swinging bucket rotor: 3,800 g/ 4,800 rpm

6. Clinical Centrifuge • Uses: Serum, urine and blood sedimentation • Fixed angle rotor and swinging bucket: 4600 g/ 6000 rpm

7. Microfuge • Uses: Cells (at low speeds) and to spin down a reagent in a tube • Fixed angle (rare swinging bucket): 16,000 g/ 13,000 rpm

8. High Speed Centrifuge • Uses: Large volume, pelleting bacteria, spin columns and protein precipitations • Fixed angle rotor and swinging buckets: 82,000 g / 26,000 rpm

9. Ultracentrifuge • Uses: Virus concentration, membrane and subcellular fraction isolation, DNA and RNA isolation • Fixed angle rotor and swinging bucket: 800,000 g / 100,000 rpm

10. Benchtop Ultracentrifuge • Uses: Membrane preps, virus isolation, subcellular fractionation, CsCl DNA plasmid isolations in 30 minutes • Fixed angle rotor and swinging bucket: 625,000 g/ 120,000 rpm

11. Rotors • The internal rotating unit within a centrifuge which holds tubes and plates. Fixed angle rotor Swinging bucket rotor

12. Primary hazards • Mechanical failure • Dispersion of aerosols • (hazardous aerosols)

13. Mechanical Failure • Is caused by age and by improper use or inadequate care of centrifuge or rotor. Especially the rotor.

14. Care and Attention • Safe high-speed spin requires nearly perfectly balanced load. • Age, use, and misuse contribute to rotor flaws. • A rotor which comes apart at high speed can be deadly.

15. Tiny flaws are …NOT so tiny at 80,000 rpm These are micro-fissures and cracks caused by stress and corrosion in the bottom of a tube cavity.

16. (Purdue 2003) This rotor came apart while coming up to speed (not yet spinning at full speed) The 6 inch (15 cm) long chunk damaged (ruined) but did not completely penetrate the centrifuge lid and housing. The motor and spindle were also destroyed.

17. To reduce wear and chance of failure • Follow instructions in manual and rotor care guide • Use only compatible rotors • Check routinely for rotor damage • Don't overfill tubes • Cap tubes • Balance load carefully • Check that rotor correctly seated on drive spindle • Run at < max safe speed continued……

18. To reduce wear and chance of failure, continued • Stay right there until full speed is reached. • Stop the centrifuge if anything seems unusual • Never open until stopped (never ‘manual brake’) • Clean rotor gently • Dry completely • Store rotor upside down (why?) • Maintain a careful rotor log • Derate and retire rotors for age/use

19. (MIT 1999) This rotor split in half at 55,000 rpm after 3 h of what was supposed to be a much longer spin. Grad student report “Rotor was manufactured in 1986 and was covered for 10,000 hours of operation or 5 years, whichever comes first.” No surprise that it flew apart 13 years later. Fortunately the centrifuge housing contained it.

20. (Cornell, 1998) • The rotor failure was not contained. • Much other damage done, equipment ruined, chemicals spilled. • Nobody was near, or someone would have been badly hurt or killed.