To carry out their activities, Research Teams of the Frédéric Joliot Institute for Life Sciences have developed high-profile technological platforms in many areas : biomedical imaging, structural biology, metabolomics, High-Throughput screening, level 3 microbiological safety laboratory...
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Scientific result | High-throughput screening | Methodologies for life sciences | Proteomics | Bacteria
Researchers at Li2D (SPI/DMTS, Marcoule) are proposing an innovative multiplexing method for rapid identification of several microbial isolates in a single mass spectrometry analysis. This approach opens up prospects for the large-scale identification of microorganisms resulting from culturomics programs.
To meet the needs of clinical diagnosis and microbiological screening in general, it is essential to be able to quickly and reliably identify the microorganisms present in a microbiota sample (normal or pathological microbial flora). In order to better understand the individual components of microbiota, culturomics is being developed as a means of generating numerous microbial isolates with highly distinct characteristics by systematically testing hundreds of different culture conditions. High-throughput identification methods now need to be developed to rapidly screen these isolates. In the present proof of concept, the researchers propose an innovative multiplex proteotyping methodology for the simultaneous analysis of 21 bacterial samples by tandem mass spectrometry, without relying on differential peptide labeling that would require costly chemical reagents. The method is based on pre-fractionation of the peptidomes of each microbial isolate by reverse-phase chromatography, mixing of specific fractions of each isolate, differentiated according to their hydrophobicity, followed by analysis of the mixture in a single nanoLC-MS/MS (Liquid Chromatography coupled to tandem Mass Spectrometry) experiment. Organism-specific signals are distinguished from other signals by their hydrophobic properties, and the process identified all 21 microorganisms in a single 60-minute analysis. The researchers also developed a specific proteotyping concept to assign taxonomic information to signals restricted by time unit, thanks to clever phylopeptidomic computer processing.
Conclusion: By developing a label-free multiplexing strategy for the analysis of microbial isolates by tandem mass spectrometry, and in view of recent technological advances in this approach, the authors of this study envisage being able to identify up to 200 microbial isolates per hour, enough to revolutionize "culturomics" and advance our knowledge of microbiota. Contacts : Béatrice Alpha-Bazin (email@example.com) – Jean Armengaud (firstname.lastname@example.org)
Madisson Chabas, Olivier Pible, Jean Armengaud, Béatrice Alpha-Bazin. Label-Free Multiplex Proteotyping of Microbial Isolates. Anal. Chem. 2023, 95, 35, 13163-13171 https://doi.org/10.1021/acs.analchem.3c01975
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