The Kerr Lab

Safety Note: Rifampicin (MSDS) is toxic and should be handled carefully. Personal protection (safety glasses, lab coat and gloves) should be worn when working with Rifampicin. All work with rifampicin should be done in the fume hood. DMF (MSDS) is also an irritant and should be handled carefully.

Materials: Gather to have under or near the hood:

• 3 beakers
  • Rifampicin waste beaker (large)
  • Beaker for dissolving rifampicin (can be much larger than desired volume)
  • Beaker for pouring DMF
• Graduated cylinder
• Spatula
• Stir bar
• Waste bag and container for disposable rifampicin waste
• Several pieces of parafilm for covering containers with rifampicin/ DMF
• 0.22 uM filter for sterilization
• Syringe of desired volume for sterilization
• 30-mL milk bottle wrapped in foil
• Rifampicin
• Fine-balance scale
• Stir plate (plug into the socket on the hood)
• Kim wipes
• Spray bottle of 70% ethanol
• Extra aluminum foil
• Extra gloves

Protocol:

1. Begin by placing the scale in the fume hood. Level the scale by adjusting the back legs until the bubble is centered.
2. Pour DMF into large beaker.
3. Pour correct amount from large beaker into graduated cylinder.
4. Cover cylinder with parafilm.
5. Put on safety gear: labcoats, masks, goggles, gloves.
6. Tare scale with small (~40ml) beaker on it, with stir bar.
7. Weigh out rifampicin powder.
   1. Keep open rifampicin bottle inside of scale doors.
   2. After touching rifampicin, use only the backs of your hands to open and close scale doors.
   3. Avoid tapping spatula on the edge of the beaker as that can mess up the scale.
   4. After weighing, put spatula in waste beaker and immediately spray down with EtOH.
   5. When done handling the powder change gloves and discard old gloves in biohazard waste.
8. After rifampicin is weighed out, note the weight in notebook and add the measured DMF to the beaker and cover with parafilm.
9. Clean scale carefully, wiping with kimwipes + EtOH until no color comes off.
10. Take scale out of fume hood.
11. Put stir plate in hood.
12. Use the stir plate to mix rifampicin and DMF. Note: Make sure the stir bar doesn’t spin too fast and knock the beaker over.
13. Once well mixed, turn the stir plate off. Double check there’s no powder clumped onto the stir bar.
14. Filter sterilize solution under the hood.
15. Wrap in tinfoil or store in a light resistant container
16. Label under foil and outside the foil with "Rifampicin in DMF, initials, date, concentration, flammable, toxic".
17. Store at 4°C.
18. Wipe down everything and put all rifampicin waste (including mixing beaker) in biohazard bin. Put foil over the rifampicin waste beaker, label it appropriately, and add it to a waste load.
19. Let DMF evaporate from the DMF beaker under the hood.
20. Note the procedure in lab notebook, including lot numbers for the rifampicin and DMF.

Recipes adapted from Miller 1992: A Short Course in Bacterial Genetics

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Orignal Recipe by: C. Eshelman Date: 6 Jul 2012

Recipe Updated by: S. Singhal Date: 6 Sep 12