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Susceptibility of the Different Oxygen-Sensing Probes to Interferences in Respirometric Bacterial Assays with Complex Media

Authors :
Chiara Zanetti
Liang Li
Rafael Di Lazaro Gaspar
Elisa Santovito
Sophia Elisseeva
Stuart G. Collins
Anita R. Maguire
Dmitri B. Papkovsky
Source :
Sensors, Vol 24, Iss 1, p 267 (2024)
Publication Year :
2024
Publisher :
MDPI AG, 2024.

Abstract

Respirometric microbial assays are gaining popularity, but their uptake is limited by the availability of optimal O2 sensing materials and the challenge of validating assays with complex real samples. We conducted a comparative evaluation of four different O2-sensing probes based on Pt-porphyrin phosphors in respirometric bacterial assays performed on standard time-resolved fluorescence reader. The macromolecular MitoXpress, nanoparticle NanO2 and small molecule PtGlc4 and PtPEG4 probes were assessed with E. coli cells in five growth media: nutrient broth (NB), McConkey (MC), Rapid Coliform ChromoSelect (RCC), M-Lauryl lauryl sulfate (MLS), and Minerals-Modified Glutamate (MMG) media. Respiration profiles of the cells were recorded and analyzed, along with densitometry profiles and quenching studies of individual media components. This revealed several limiting factors and interferences impacting assay performance, which include probe quenched lifetime, instrument temporal resolution, inner filter effects (mainly by indicator dyes), probe binding to lipophilic components, and dynamic and static quenching by media components. The study allowed for the ranking of the probes based on their ruggedness, resilience to interferences and overall performance in respirometric bacterial assays. The ‘shielded’ probe NanO2 outperformed the established MitoXpress probe and the small molecule probes PtGlc4 and PtPEG4.

Details

Language :
English
ISSN :
14248220
Volume :
24
Issue :
1
Database :
Directory of Open Access Journals
Journal :
Sensors
Publication Type :
Academic Journal
Accession number :
edsdoj.b4cddd6779a64149963a97619e5a31c4
Document Type :
article
Full Text :
https://doi.org/10.3390/s24010267