E. coli was isolated on Endo Agar (104044 - Merck Millipore, USA). As an enrichment medium, we used peptone broth supplemented with lactose and bile (Kessler-GRM medium, Obolensk, Russia). Incubation was carried out at 37°C for 24 h on both media. Identification of isolates was performed by the MALDI-TOF MS method using the MALDI Biotyper Microflex system (Bruker, USA), according to the Maldi Biotyper 3.0 User Manual. The identification results were verified by the API20E biochemical kit (BioMerieux, France). One isolate per each sample was obtained (n=306).
Maldi biotyper microflex system
Manufactured by Bruker
Sourced in Germany
The MALDI Biotyper Microflex system is a matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry platform used for microbial identification. The system generates characteristic protein profiles of microorganisms, which are then compared to a comprehensive reference database for rapid and reliable identification.
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4 protocols using maldi biotyper microflex system
1
Isolation and Identification of E. coli
2
Microbial Identification Using MALDI-TOF MS
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The obtained isolates were identified with a MALDI Biotyper Microflex system (Bruker Daltonics, Bremen, Germany) according to the manufacturer’s instructions. The protein extraction method was applied using ethanol/formic acid sample preparation [14 (link)]. 1 µL of the respective protein extract of each isolated colony was added to a spot on the Biotyper steel target plate. After air drying, the samples were overlayed with 1 μL MALDI-matrix solution (alpha-Cyano-4-hydroxycinnamic acid, Bruker Daltonics, Bremen, Germany). After further air drying, the samples were analyzed. The obtained mass spectra were compared against the internal MALDI Biotyper reference libraries: MBT Compass Library, revision F, v. 9, containing 8468 main spectra (MSPs); MBT Filamentous Fungi Library (revision No. 2, containing 468 MSPs); MBT Security Related Library (SR Library, revision No. 1; containing 104 MSPs). Matches with the respective spectra in the databases were displayed as scores ranging from 0.0 to 3.0. Scores ≥ 1.7 indicated a secure genus identification and scores ≥ 2.0 a secure genus and probable species identification [15 (link)].
3
Isolation and Identification of Enterococci
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Bacteria were isolated using chromogenic broth as a liquid enrichment medium (HiCrome enterococci broth, HiMedia, India). Slanetz-Bartley agar was used to obtain individual colonies of enterococci (Slanetz-Bartley agar HiMedia). The samples were incubated at 37°C for 24 h on both media. Identification was performed on a time-of-flight mass spectrometer (MALDI Biotyper Microflex system, Bruker, Germany). The identification results were confirmed by the API® Strep biochemical kit (BioMerieux, France).
4
Identification of isolates by MALDI Biotyping
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The obtained isolates were identified with a MALDI Biotyper Microflex system (Bruker Daltonics, Bremen, Germany) according to the manufacturer's instructions. The protein preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
extraction method was applied using ethanol/formic acid sample preparation. 1 µl of the respective protein extract of each isolated colony was added to a spot on the Biotyper steel target plate. After air drying, the samples were overlayed with 1 l MALDI-matrix solution (alpha-Cyano-4-hydroxycinnamic acid, Bruker Daltonics). After further air drying, the samples were analyzed. The obtained mass spectra were compared against the internal MALDI Biotyper reference libraries: MBT Compass Library, revision F, version 9, containing 8468 main spectra (MSPs); MBT Filamentous Fungi Library (revision No. 2, containing 468 MSPs); MBT Security Related Library (SR Library, revision No. 1; containing 104 MSPs). Matches with the respective spectra in the databases were displayed as scores ranging from 0.0 -3.0. Scores 1.7 indicated a secure genus identification, scores 2.0 a secure genus and probable species identification (12) .
extraction method was applied using ethanol/formic acid sample preparation. 1 µl of the respective protein extract of each isolated colony was added to a spot on the Biotyper steel target plate. After air drying, the samples were overlayed with 1 l MALDI-matrix solution (alpha-Cyano-4-hydroxycinnamic acid, Bruker Daltonics). After further air drying, the samples were analyzed. The obtained mass spectra were compared against the internal MALDI Biotyper reference libraries: MBT Compass Library, revision F, version 9, containing 8468 main spectra (MSPs); MBT Filamentous Fungi Library (revision No. 2, containing 468 MSPs); MBT Security Related Library (SR Library, revision No. 1; containing 104 MSPs). Matches with the respective spectra in the databases were displayed as scores ranging from 0.0 -3.0. Scores 1.7 indicated a secure genus identification, scores 2.0 a secure genus and probable species identification (12) .
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