MALDI-TOF MS analysis was conducted on a Bruker UltrafleXtreme MALDI-TOF/TOF MS instrument (Ultraflextreme, Bruker, Germany) operated in reflection-positive ion mode with an acceleration voltage of 20 kV. The sample was mixed with matrix in a 1:1 (v/v) ratio. The matrix was α-cyano-4-hydroxycinnamic acid, which was dissolved in 50% acetonitrile containing 0.1% trifluoroacetic acid (TFA) (Monti et al., 2011 (link)). The nitrogen laser was set at a threshold adequate for signal generation that minimized fragmentation.
Ultraflextreme maldi tof tof ms instrument
Manufactured by Bruker
Sourced in Germany
The UltrafleXtreme MALDI-TOF/TOF MS instrument is a high-performance mass spectrometry system designed for protein and peptide analysis. It combines matrix-assisted laser desorption/ionization (MALDI) technology with tandem time-of-flight (TOF/TOF) mass analysis to provide accurate and sensitive mass measurements of samples.
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4 protocols using ultraflextreme maldi tof tof ms instrument
1
MALDI-TOF MS Protein Analysis Protocol
2
MALDI-TOF MS Identification of Equine Wound Isolates
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MALDI-TOF MS was performed for the identification of isolated microorganisms from horse wounds as previously described [12 , 44 (link)] with few modifications. Bacterial isolates were cultured as described above and the direct transfer formic acid method was used for all samples [44 (link)]. The experiments were performed in linear-positive mode on Ultraflextreme MALDI-TOF/TOF MS instrument (Bruker, Sweden) in a mass range of 2–20 kDa. Mass spectra were analyzed using the FlexControl and MALDI Biotyper 3.1 software with the BDAL-5627 reference database (Bruker Daltronics, Sweden). Samples that were not identified by MALDI-TOF MS were prepared for 16S rRNA gene polymerase chain reaction (PCR) amplification and sequencing.
3
MALDI-MS Protein Identification Protocol
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MALDI-MS mass spectra were recorded in a positive ion measurement mode using Ultraflextreme MALDI-ToF-ToF-MS instrument (Bruker Daltonics, Germany). The spectra were obtained in reflecto-mode with the accuracy of measuring the monoisotopic m/z ratio up to 0.1 Da. The fragmentation spectra were obtained using the Lift mode, for which the accuracy of measuring the daughter ions was within 0.2 Da. Sample aliquots were mixed on a steel target with a 30 mg/ml 2,5-dihydroxybenzoic acid in 0.5% TFA and a 30% MeCN–water solution (Sigma-Aldrich) (see Supplementary Data 1 –4 ).
4
MALDI-TOF/TOF MS Tissue Calibration
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Firstly, red phosphorous (Sigma-Aldrich, Castle Hill, Australia) was mixed with methanol (1 mg/mL) and manually spotted (1 μL) onto each flatworm to externally calibrate the UltrafleXtreme MALDI-TOF/TOF MS instrument (Bruker Daltonics, Bremen, Germany). Slides were marked at the edges with water-based white out, and then scanned at 4800 dpi on a CanoScan 5600F (Canon Australia, Macquarie Park, Australia), scanner to teach the instrument the spatial orientation of the slide in relation to the laser. Subsequently, α-cyano-4-hydroxycinnamic acid (CHCA; 7 mg/mL in 50% acetonitrile, 0.2% v/v Trifluoroacetic acid) was deposited onto each slide using an iMatrixSpray, Subingen, Switzerland, instrument, and the following instrument-specific settings: 6 mm height with 1 mm line distance. Instrument speed was at 160 mm/s. The matrix cover was 1 μL/cm2 (9.8 units/cm2) density, 30 cycles, 15 s delay, and 80 × 30 mm dimensions.
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