Bdellovibrio sp.

1. Fill 13 mm screw cap test tubes completely to the top with #550 broth and then add 0.02 mL of 3% cysteine.
2. Let the test tubes sit for at least one hour prior to opening.
3. Open vial according to enclosed instructions.
4. Using a single tube of #550 reduced broth (5 to 6 mL), withdraw approximately 0.5 to 1.0 mL with a Pasteur or mL pipette. Rehydrate the entire pellet.
5. Aseptically transfer this aliquot back into the broth tube. Mix well.
6. Use 0.5 mL of the suspension to inoculate a #550 secondary broth tube.
7. Incubate the tubes at 30°C for 3 to 7 days under a tungsten lamp. Ensure that the screw cap tops are sealed during the incubation period.

1. Open a vial of ATCC^® 27047™ according to enclosed instructions.
2. Rehydrate the entire pellet with 0.5 mL of #587 broth. Withdraw suspension into a sterile test-tube.
3. Add approximately 0.5 mL of the host strain.             
4. Melt tubes containing 2.5 mL of 0.8% semi-solid agar, and place in a 45°C water bath.
5. Add 0.2 to 0.3 mL of the host/predator suspension to each melted tube and pour over the surface of pre-warmed #587 plates.
6. Incubate plates anaerobically at 30°C for at least 10 days.  After this time, plaques may be visible.  Observe daily until plaques or clearing are seen. 
7. Scrape off overlay into a test-tube and centrifuge at low speed to sediment the agar and cellular debris.  The Bdellovibrio cells should be in the supernatant. Check for tiny, rapidly moving cells under a wet mount.

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