Chapter 5
Detection of the Bacterial Quorum-Sensing
Signaling MoleculesN-Acyl-Homoserine Lactones
(HSL) andN-Acyl-Homoserine (HS) with an Enzyme-
Linked Immunosorbent Assay (ELISA) and via
Ultrahigh-Performance Liquid Chromatography
Coupled to Mass Spectrometry (UHPLC-MS)
Michael Rothballer, Jenny Uhl, Josie Kunze, Philippe Schmitt-Kopplin,
and Anton Hartmann
Abstract
Quick and reliable quantitative methods requiring low amounts of sample volume are needed for the
detection ofN-acyl-homoserine lactones (HSL) and their degradation productsN-acyl-homoserines (HS)
in order to elucidate the occurrence and dynamics of these prevalent quorum-sensing molecules of Gram-
negative bacteria in natural samples and laboratory model experiments. A combination of ELISA and
UHPLC-MS is presented here which has proven to meet these requirements. Both methods can not only
precisely detect and quantify HSLs but also their degradation products HS and thereby enable studying
signaling dynamics in quorum sensing, which have been identified to play an essential role in bacterial
communication.
Key wordsN-acyl-homoserine lactone, Quorum sensing, Bacterial signaling, ELISA, UHPLC-MS1 Introduction
For the analysis of quorum-sensing (QS) signaling in Gram-
negative bacteria it is mandatory to measure exactly what kind of
signaling substances are present in a certain habitat and at what
concentration. For the best known QS-signaling substance,N-acyl-
homoserine lactones (HSLs), several detection methods have been
developed, e.g., biosensor assays, thin-layer chromatography, and
gas and liquid chromatography without or coupled with mass
spectrometry [1–7]. All these methods have certain advantages
and drawbacks, including difficulties in quantification of autoindu-
cers produced at low level, although low concentrations of signalLivia Leoni and Giordano Rampioni (eds.),Quorum Sensing: Methods and Protocols, Methods in Molecular Biology,
vol. 1673,https://doi.org/10.1007/978-1-4939-7309-5_5,©Springer Science+Business Media LLC 2018
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