Summary

In this tutorial you learned how to dissolve a Vitroids™ disc and prepare the bacterial suspension for BactoBox® analyses.

Calculate cell concentration of non-diluted suspension

You just did your first BactoBox® measurement. The result on the BactoBox® is for the diluted sample. Always remember to multiply the obtained result with the dilution factor to get the concentration of the non-diluted suspension.

BactoBox® dilution	BactoBox® concentration	Non-diluted suspension
100	89 000 cells/mL	100 × 89 000 cells/mL = 8 900 000 cells/mL

Comparison between CFU and BactoBox® results

We can estimate the number of cells in the Vitroids™ disc from the measured cell concentration after dissolution. We assume that the total volume remains 110 µL (0.110 ml) after the disc is added. That gives a volume of 0.110 ml/disc.

1. Measure the cell concentration in the dissolved sample: 8,900,000 cells/mL
2. Multiply by the dissolution volume: 8,900,000 cells/mL × 0.110 mL/disc = 979,000 cells/disc

To develop this tutorial, we repeated the procedure with three Vitroids™ discs from the same lot. The table below summarizes the results (all values are reported as cells/disc).

#1	#2	#3	Arithmetic mean ± Standard deviation
979 000	970 000	1 100 000	1 000 000 ± 90 000 cells/disc

The concentration is stated by Sigma/Merck as 50,000-150,000 CFU mean value range. This is ~1 log below the results obtained with the BactoBox® measurements. It is normal to see cells ≠ CFU for formulated, stored products, see the box for more information.

BactoBox detects all forms of bacteria. Plate counts only detects culturable cells.

A plate count only measures the culturable cells and does not detect VBNC forms of bacteria. BactoBox® detects viable, dead and VBNC forms of bacteria. BactoBox® shows great correlation for most Active cultures while BactoBox® measurements on bacteria in dormant and stressed states may show Divergence from plate counts.

BactoBox® skills

Congratulations! After successfully completing this tutorial you have acquired the following skills:

Disaggregrate bacterial clumps by vortexing

Make a 1:100 dilution of a dilute bacterial sample and measure on BactoBox®

Calculate cell concentrations of the non-diluted bacterial suspension

Next steps

This, first tutorial was a quick and simple introduction to a bacterial measurements.

We have another great tutorial lined up for you: Measure a dense culture. You will prepare a dense culture using a Vitroids™ disc (or an in-house E. coli cryo stock). After overnight incubation you will learn how to dilute the sample 1:10,000 to hit the right concentration.