Detection and Inhibition of Bacterial Cell–Cell Communication - Bacterial Pathogenesis: Methods and Protocols - page 63

7. This method will not identify the chemical species that induces or inhibits the
quorum-sensing monitor strain. There are other methods that can aid in this,
such as thin layer chromatography or gas chromatography-mass spectroscopy
methods (described in ref. (19) ).
8. The timing of addition relative to sampling is partly dependent on the stability
of the signal or antagonist in the presence of bacteria. Therefore, if no effect
is observed with a single dose, it may be necessary to repeat the dose during
growth or to time the addition of the extract more closely with the onset of
elastase production.
9. Filtration typically results in the loss of some sample volume, so a slight excess
is collected.
10. Normalization for cell density can be done by dividing the OD 495 by either the
OD 600 or by protein concentration.
11. It is important to note that 5 g/L of glucose has been shown to inhibit biolu-
minescence in V. harveyi (25) ; thus care must be taken to ensure the growth
medium used for the test strains does not contain high levels glucose. The cell-
free supernatant can be made glucose depleted by growing V. harveyi strain
MM30 (AI-2 − ) in the supernatant prior to testing (26) . By contrast, low levels
of glucose may actually increase bioluminescence and lead to false positives.
12. It is important that the cell-free supernatants be neutral in pH as acidity can
interfere with the assay.

Acknowledgments
This work has been supported by the National Health and Medical Research
Council of Australia and funding from Biosignal Ltd, Australia.

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