Lipid A was precipitated from GMMA as previously described [25 (link)] using mild acid hydrolysis with 1% acetic acid for 2 h at 100°C. Samples were centrifuged at 14,000 g for 15 min, the pellets resuspended in water, and washed twice with water. The pellets were dried overnight using a Speedvac, resuspended in chloroform-methanol 4:1, and mixed with an equal volume of Super DHB (Sigma-Aldrich) solution in water/acetonitrile 1:1 (vol/vol). Two μL of the mixture were loaded to the target plate (MTP 384 target plate ground steel BC, Bruker Daltonics) and analyzed by Ultraflex MALDI-TOF (Bruker Daltonics) in reflectron ion-negative mode. A Peptide Calibration Standard (Bruker Daltonics), mixed with the Super DHB solution, was included in each analysis. The m/z rations were determined by Flex Analysis software in comparison to the Peptide Standard.
Mtp 384 target plate ground steel bc
Manufactured by Bruker
Sourced in Germany
The MTP 384 target plate ground steel BC is a laboratory equipment product offered by Bruker. It serves as a target plate for various analytical applications. The product is made of ground steel and is designed to accommodate 384 sample positions.
Automatically generated - may contain errors
Lab products found in correlation
Ultraflex maldi tof tof instrument, by Bruker (2 mentions)
Peptide calibration standard, by Bruker (1 mentions)
Super dhb, by Bruker (1 mentions)
Super dhb, by Merck Group (1 mentions)
Speedvac, by Merck Group (1 mentions)
Super dhb solution, by Merck Group (1 mentions)
Lift system, by Bruker (1 mentions)
Cellulose acetate filter, by Sartorius (1 mentions)
7 protocols using mtp 384 target plate ground steel bc
1
Isolation and Characterization of Lipid A from GMMA
2
Lipid A Extraction and Analysis
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Lipid A was precipitated from GMMA using mild acid hydrolysis with 1% acetic acid for 2 h at 100 °C (35 ). Samples were centrifuged at 14,000 × g for 15 min; the pellets were resuspended in water and washed twice with water. The pellets were dried overnight using a SpeedVac and resuspended in chloroform/methanol 4:1 and mixed with an equal volume of Super DHB solution (Sigma). 2 μl of the mixture were loaded to the target plate (MTP 384 target plate ground steel BC, Bruker Daltonics) and analyzed by Ultraflex MALDI-TOF (Bruker Daltonics) in reflectron ion-negative mode. A peptide calibration standard (Bruker Daltonics), mixed with the Super DHB solution, was included in each analysis. For MS/MS analysis of lipid A, main peaks from the linear mode analysis were selected for collision-induced dissociation, and the resulting fragments were detected by MALDI TOF-TOF in ion negative mode. For each sample, spectra represent the integration of the analysis of 20 different areas of the spot by 50 single laser shots. The m/z rations were determined by Flex Analysis software in comparison with the peptide standard.
3
MALDI-TOF/TOF MS Analysis of LFCs
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The molecular weights of LFCs were confirmed by MALDI-TOF/TOF MS using an Autoflex maX TOF/TOF mass spectrometer with a laser frequency of 200 Hz (Bruker Daltonics, Billerica, MA, USA). α-Cyano-4-hydroxycinnamic acid (CHCA, MW 189.04 Da) matrix was chosen as the peptide calibration reagent with MW <10,000 Da. In addition, 2,5-dihydroxybenzoic acid (2,5-DHB, MW 154.03 Da) was used for our LFC analysis. After Prep-HPLC purification, the LFC sample was dissolved in 75% ACN solution, and a mixture of 2 μL samples and 2 μL 2.5-DHB matrix was prepared in a 1:1 ratio. The sample was loaded into the template (MTP 384 target plate ground steel BC, Bruker) and then measured after drying the organic solvent with a vacuum dryer.
4
MALDI-TOF Analysis of Oligosaccharides
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One μL sample (arabino-, fructo-, or galacto-oligosaccharides, 10 mg/mL oligosaccharide dissolved in MilliQ water) and 1 μL of 10 mg/mL di-hydroxy benzoic acid matrix in 50% acetonitrile was applied to a MTP 384 target plate ground steel BC (Bruker Daltonics GmbH, Bremen, Germany), mixed on the target and dried under a stream of air. The samples were analyzed with an Ultraflex MALDI-TOF/TOF instrument (Bruker Daltonics GmbH, Bremen, Germany). The instrument was operated in positive acquisition mode and controlled by the FlexControl 3.3 software package. All spectra were obtained in reflector mode with the least required laser intensity and pulsed ion extraction of 40 ns. The acquisition range used was from m/z 300 to 3000.
5
MALDI-TOF/TOF Mass Spectrometry Protocol
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A 1 μL sample and 1 μL of 10 mg/mL norharmane matrix in 50% acetonitrile was applied to a MTP 384 target plate ground steel BC (Bruker Daltonics GmbH, Bremen, Germany), mixed on the target and dried under a stream of air. The samples were analyzed with an Ultraflex MALDI-TOF/TOF instrument (Bruker Daltonics GmbH, Bremen, Germany). The instrument was operated in negative acquisition mode and controlled by the FlexControl 3.3 software package. All spectra were obtained using the linear mode with an acceleration voltage of 20 kV, and pulsed ion extraction of 10 ns. The acquisition range used was from m/z 400 to 2500. Post-source decay (PSD) spectra using the Bruker Daltonics LIFT system were recorded at 8 kV precursor ion acceleration voltage and fragment acceleration (LIFT voltage 19 kV). The reflector voltage 1 and 2 were set to 29.6 and 13.9 kV, respectively.
6
Soil Extract Analysis by LDI-TOF MS
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Ten grams of homogenized fresh soil sample from 0 to 4 cm soil depth of the control and the grasshopper mesocosms was each weighted into a 45-mL centrifuge tube. Thirty milliliters of deionized water was added and horizontally shaken for 2 h at 150 rpm/min. After 5 min of centrifugation at 2800×g, the solutions were filtered under vacuum (0.45 μm pore size, cellulose acetate filter, Sartorius, Göttingen, Germany) and the resulting extracts were freeze-dried immediately. Subsequently, the extracts were re-suspended in 500 μL methanol and 1 μL was spotted on a MTP 384 target plate ground steel BC (Bruker Daltonik). The LDI-TOF MS experiments were performed as well on an UltrafleXtreme MALDI-TOF/TOF mass spectrometer in the positive ion mode within a mass range of m/z 150 to 2000. The resulting mass spectra are presented in Fig. 7 .
7
MALDI-TOF Sample Preparation Protocol
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A small amount (one tip of a wooden application stick) of one colony was transferred directly from the culture medium onto a ground steel target (MTP 384 target plate ground steel BC, Bruker Daltonics GmbH, Bremen, Germany). The spot was overlaid with 1 μl matrix and air-dried at room temperature (approx. 22 °C). After the spot was dried, the sample was again overlaid by 1 μl matrix and allowed to air-dry at room temperature.
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