Use 1:10 000 dilutions in deceleration and stationary stage

Switch to using a 1:10,000 dilution when the 1:1,000 dilution yields more than 1,500,000 total/ml for the last sample.

Prepare a 1:10 000 dilution

1. Use a serological pipette to collect a sample from the shake flask.

2. Return the shake flask to the incubator shaker.

3. Start a 30-minute countdown to remind you to collect the next sample.

4. Vortex 30 seconds at max speed to disaggregate bacteria.

5. Make a 1:100 dilution:

   1. Transfer 101 µL of your sample to 10 mL of diluent.

   2. Vortex 10 seconds at maximum speed.

6. Make a 1:10 000 dilution:

   1. Transfer 101 µL of the 1:100 dilution to 10 mL diluent.

   2. Vortex 10 seconds at maximum speed.

7. Transfer the tubing kit to the 1:10 000 vial.

Do a Clean program after each measurement.

Start a 30-minute countdown to remind you to collect the next sample.

Measure the diluted samples in Access

1. Click the Measure button in Access.

2. Add metadata:

   1. Dilution: 10 000.

   2. Label: E_coli_growth.

Clean the device

1. Transfer the tubing kit to the disinfection vial.

2. Click Clean.

Summary

You have now tracked the concentration through lag, exponential, deceleration and stationary stage. The final step is to determine when to conclude the growth curve experiment.