Ultraflex 2 maldi tof tof mass spectrometer

Manufactured by Bruker

Sourced in Germany, United States

The Ultraflex II MALDI-TOF/TOF mass spectrometer is a high-performance analytical instrument designed for the characterization of complex biomolecules. It combines matrix-assisted laser desorption/ionization (MALDI) with time-of-flight (TOF) mass spectrometry and tandem TOF (TOF/TOF) capabilities to provide accurate mass determination and structural analysis of various samples, including proteins, peptides, and small molecules.

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Lab products found in correlation

Flexanalysis 3, by Bruker (3 mentions) 2 5 dihydroxybenzoic acid, by Merck Group (2 mentions) Biotools 3, by Bruker (2 mentions) Pepmap c18, by Thermo Fisher Scientific (1 mentions) Ulc ms grade water, by Biosolve (1 mentions) Acetonitrile, by Merck Group (1 mentions) Trifluoroacetic acid, by Merck Group (1 mentions) Sinapinic acid, by Merck Group (1 mentions) Sinapinic acid matrix solution, by Bruker (1 mentions) Trypsin solution, by Promega (1 mentions) Bacterial test standard, by Bruker (1 mentions) Geloader tip, by Eppendorf (1 mentions) Iodoacetamide, by MP Biomedicals (1 mentions) Iodoacetamide, by Promega (1 mentions) Trypsin, by Promega (1 mentions) Sequencing grade trypsin, by Promega (1 mentions) Mascot software, by Matrix Science (1 mentions) Methanol, by Bruker (1 mentions) Fraction 5, by Merck Group (1 mentions) Amazon sl quadrupole ion trap mass spectrometer, by Bruker (1 mentions)

Spelling variants of this product

MALDI-TOF mass spectrometer (57 citations) MALDI-TOF/TOF mass spectrometer (45 citations) Ultraflex MALDI-TOF mass spectrometer (31 citations) Ultraflex MALDI-TOF-TOF mass spectrometer (26 citations) Ultraflex II MALDI-TOF/TOF (18 citations) Ultraflex MALDI-TOF/TOF (17 citations) Ultraflex II mass spectrometer (13 citations) UltraFlex II MALDI-TOF Mass Spectrometer (12 citations) Ultraflex II TOF/TOF (12 citations) Ultraflex II MALDI mass spectrometer (2 citations)

13 protocols using ultraflex 2 maldi tof tof mass spectrometer

Peptide Separation and Identification

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Samples were dissolved in 15 µL of 5% ACN in water and filtered. Ten microliters of the sample were loaded onto the guard column with an isocratic flow of 2% ACN in ULC/MS grade water (Biosolve, Lexington, MA) and 0.1% FA, at a flow rate of 10 µL/min. After 2 min, the guard column was switched online with a PepMap C18, 3 µm × 150 mm nanocolumn (Thermo Fisher Scientific) analytical capillary column. Separation was conducted using a linear gradient from 95% solvent A, 5% solvent B to 5% solvent A, 65% solvent B in 35 min (solvent A: H₂0/ACN/FA (94.9:5:0.1, v/v/v); solvent B: ACN/FA (99.9:0.1, v/v)), at a flow rate of 200 nL/min.
Fractions were collected manually using 15 s intervals on a ground steel MALDI target plate (MTP 384, Bruker Daltonics). Prior to fraction collection, the sample plate was prespotted with α-cyano-4-hydroxycinnamic acid (CHCA) using 0.5 µl of a 5× dilution of a saturated solution of CHCA in 100% acetone. Fractions were collected in the center of each target spot within 10 to 35 min of the LC separation and left to dry, and then analyzed with the ultraflex II MALDI-TOF/TOF mass spectrometer (Bruker Daltonics). Mass spectra were acquired in reflectron mode from m/z 750 to 4000. Ions of sufficient intensity were selected manually for fragmentation analysis.

Monroe E.B., Annangudi S.P., Wadhams A.A., Richmond T.A., Yang N., Southey B.R., Romanova E.V., Schoofs L., Baggerman G, & Sweedler J.V. (2018). Exploring the sea urchin neuropeptide landscape by mass spectrometry. Journal of the American Society for Mass Spectrometry, 29(5), 923-934.

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Mass Spectrometric Analysis of Phosphotyrosine Intermediate

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To prove the covalent phosphotyrosine intermediate formation for Ser35Arg, we performed mass spectrometric analysis of the tryptic hydrolysate of the Ser35Arg mutant before and after interaction with paraoxon. The Ser35Arg antibody (10 μM) was incubated with paraoxon [2 mM in 0.1 M phosphate buffer (pH 7.4)] at 37°C for 16 hours. Samples were treated with sequencing-grade trypsin according to the Bruker applications guide and analyzed directly using UltraFlex II MALDI-TOF/TOF mass spectrometer (Bruker Daltonics). Analysis was performed with flexAnalysis 3.3 (Bruker Daltonics). The mass difference (M/e, 136) of the multicharged peaks corresponded to the mass of a diethoxyphosphoryl residue (fig. S9).

Smirnov I.V., Golovin A.V., Chatziefthimiou S.D., Stepanova A.V., Peng Y., Zolotareva O.I., Belogurov AA J.r., Kurkova I.N., Ponomarenko N.A., Wilmanns M., Blackburn G.M., Gabibov A.G, & Lerner R.A. (2016). Robotic QM/MM-driven maturation of antibody combining sites. Science Advances, 2(10), e1501695.

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MALDI-TOF-TOF Protein Identification

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Protein spots with significant differences between the two groups were excised, dehydrated in acetonitrile, and dried at room temperature. Gel pieces were denatured, alkylated, trypsin digested and analyzed by an Ultraflex II MALDI-TOF-TOF mass spectrometer (Bruker Daltonics GmbH, Bremen, Germany) under the control of FlexControl TM 2.4 software (Bruker Daltonics GmbH). Acquired peptide mass fingerprint (PMF) were processed using the software FlexAnalysis™ 3.0 (Bruker Daltonics, Bremen, Germany). The peak detection algorithm was: SNAP (Sort Neaten Assign and Place); S/N threshold: 1.5; Quality Factor Threshold: 50. The tryptic auto-digestion ion picks (trypsin [108–115] 842.5094 Da, trypsin [58–77] 2211.104 Da) were used as internal standards. The resulting peptide mass lists were used to search the Matrixscience database (http://www.matrixscience.com). The following search parameter criteria were used: mass tolerance 100 ppm, miss cleavage≦1, modification comprises carbamidomethyl and methionine oxidation. Matched peptides number between experimental PMF and theoretical PMF≧5 [58 (link)]. All MS data have been deposited in PeptideAtlas and are accessible through Dataset Identifier PASS00597 (http://www.peptideatlas.org/PASS/PASS00597).

Liu N., Li H., Liu K., Yu J., Bu R., Cheng M., De W., Liu J., He G, & Zhao J. (2014). Identification of skin-expressed genes possibly associated with wool growth regulation of Aohan fine wool sheep. BMC Genetics, 15, 144.

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Leptospira MALDI-TOF Mass Spectrometry

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Leptospira was cultured for a week to obtain 10⁸ colony forming unit (CFU) per ml as previously described [14 (link)]. Three milliliters of culture were centrifuged at 13,000 g for 2 min at room temperature and cell pellet was washed with 70% ethanol and centrifuged at 13,000 g for 2 min. The pellet was mixed with 50 μl sinapinic acid matrix solution (Bruker Daltonics, Germany) containing 10 mg of sinapinic acid in 1 ml of 2.5% trifluoroacetic acid (Sigma-Aldrich, USA) and 50% acetonitrile (Sigma-Aldrich, USA). After being well mixed by pipetting and centrifuged at 13,000 g for 1 min, then 2 μl of cell-matrix suspension were added to the ground steel MALDI target plate and dried at room temperature. Each isolates were spotted 24 dots onto the MALDI target plate to test technical replication. Mass spectra were collected using a Ultraflex II MALDI-TOF/TOF mass spectrometer (Bruker Daltonic, USA) operated with FlexControl software in linear positive mode, i.e. using a mass range of 2,000 to 20,000 Daltons. The instrument was externally calibrated with E. coli strain DH5α ribosomal proteins as recommended protocol.

Sonthayanon P., Jaresitthikunchai J., Mangmee S., Thiangtrongjit T., Wuthiekanun V., Amornchai P., Newton P., Phetsouvanh R., Day N.P, & Roytrakul S. (2019). Whole cell matrix assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for identification of Leptospira spp. in Thailand and Lao PDR. PLoS Neglected Tropical Diseases, 13(4), e0007232.

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Proteomic Identification of Differentially Expressed Proteins

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Protein spots with significant differences between the two groups were excised, dehydrated in acetonitrile, and dried at room temperature^{28 (link)}. Gel pieces were denatured, alkylated, trypsin digested and analysed by an Ultraflex II MALDI-TOF-TOF mass spectrometer (Bruker Daltonics GmbH, Bremen, Germany) under the control of FlexControl TM 2.4 software (Bruker Daltonics GmbH)^{28 (link)}. Acquired peptide mass fingerprint (PMF) were processed using the software Flex Analysis^TM 3.0 (Bruker Daltonics, Bremen, Germany)^{28 (link)}. The peak detection algorithm was: SNAP (Sort Neaten Assign and Place); S/N threshold: 1.5; Quality Factor Threshold: 50. The tryptic auto-digestion ion picks (trypsin [108–115] 842.5094 Da, trypsin [58–77] 2211.104 Da) were used as internal standards^{28 (link)}. The resulting peptide mass lists were used to search the Matrix science database (http://www.matrixscience.com)^{28 (link)}. The following search parameter criteria were used^{65 (link)}: mass tolerance 100 ppm, miss cleavage ≤ 1, modification comprises Carbamidomethyl and methionine oxidation^{28 (link)}. Matched peptides number between experimental PMF and theoretical PMF ≥ 5^{28 (link)}.

Zhao J., Qin H., Xin J., Liu N., Han R., Perez-Campo F.M, & Li H. (2020). Discovery of genes and proteins possibly regulating mean wool fibre diameter using cDNA microarray and proteomic approaches. Scientific Reports, 10, 7726.

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2D DIGE Protein Identification via MALDI-TOF Mass Spectrometry

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The protein spots (after 2D DIGE) were subjected to trypsin in-gel hydrolysis as described [46 (link)]. Gel pieces of 2 mm³ were excised and washed with 10 μL of a 15 mM sodium thiosulfate and 50 mM potassium hexacyanoferrate (III) mixture for 10 min at room temperature, washed twice with deionized water, dehydrated with 100 μL of acetonitrile, and air-dried. The gel pieces were subsequently treated with 3 μL of a 15 mg/mL trypsin solution (Promega, Madison, WI, USA) in 50 mM ammonium bicarbonate for 16 h at 37 °C. Peptides were extracted with a 0.5% trifluoroacetic acid water solution (6 μL) for 30 min.
Aliquots (2 μL) of the sample were mixed on a ground steel target with a 0.5 μL of 2,5-dihydroxybenzoic acid (Sigma-Aldrich, Steinheim, Germany) solution (30 mg/mL in 30% acetonitrile/ 0.5% trifluoroacetic acid), and the droplet was left to dry at room temperature. Mass spectra were recorded on an Ultraflex II MALDI-ToF-ToF mass spectrometer (Bruker Daltonik, Bremen, Germany) equipped with Nd laser. The [M + H]⁺ molecular ions were measured in a reflector mode; the accuracy of mass peak measurement was 70 ppm.

Panyushkina A., Matyushkina D, & Pobeguts O. (2020). Understanding Stress Response to High-Arsenic Gold-Bearing Sulfide Concentrate in Extremely Metal-Resistant Acidophile Sulfobacillus thermotolerans. Microorganisms, 8(7), 1076.

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MALDI-TOF/TOF Mass Spectrometry Protocol

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For database construction and sample identification, measurements were performed in AutoExecute mode using an UltraFlex II MALDI TOF/TOF mass spectrometer (Bruker Daltonics GmbH, Leipzig, Germany) with ion source 1: 25 kV; ion source 2: 23.50 kV; and a 50.0 Hz nitrogen laser. Spectra were recorded in the positive linear mode for the relative molecular mass to charge ratio (m/z) range of 2000–20,000. Each spectrum was obtained by averaging 600 laser shots. The spectra were externally calibrated by using a Bacterial Test Standard that covers a mass range of 3600–17,000 Da (Bruker Daltonics).

Emami K., Nelson A., Hack E., Zhang J., Green D.H., Caldwell G.S, & Mesbahi E. (2016). MALDI-TOF Mass Spectrometry Discriminates Known Species and Marine Environmental Isolates of Pseudoalteromonas. Frontiers in Microbiology, 7, 104.

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MALDI-TOF/TOF Mass Spectrometry of Proteins

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For MS analysis, small pieces of the gels with stained proteins of interest were cut out, incubated in 100 μl of 0.1 М NH₄HCO₃ containing 40% acetonitrile at 37°C for 20 min to remove coomassie brilliant blue, and the proteins were hydrolyzed by incubation at 37°C with 15 μg/ml solution of modified trypsin (Promega, USA) for 8 h. The solution above the gel containing the protein hydrolysate was collected and used for mass spectrometry with 2,5-dihydroxybenzoic acid (Aldrich, USA) as a matrix. Mass spectra were obtained using a Bruker Ultraflex II MALDI TOF/TOF mass spectrometer (Germany) with a reflectron.
Mass spectra data were processed using the FlexAnalysis 3.3 program (Bruker Daltonics, Germany). The search with combined data of the peptide masses and peptide fragmentation was performed by Biotools 3.2 (Bruker Daltonics). Ions score cut-off (p<0.05). Additionally, sequences of the peptides individually derived from the fragmentation data were analyzed using the T. hirsuta peroxidases dataset obtained after genome analysis.

Vasina D.V., Moiseenko K.V., Fedorova T.V, & Tyazhelova T.V. (2017). Lignin-degrading peroxidases in white-rot fungus Trametes hirsuta 072. Absolute expression quantification of full multigene family. PLoS ONE, 12(3), e0173813.

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Identification of PilG Fragments by MS

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PilG fragments selected from the endoproteinase cleavage assay were identified by peptide mass fingerprinting/MS as previously described [58 (link)]. In brief, tryptic peptides obtained from in-gel digestion were desalted and concentrated using C18 3M Empore Extraction Disks (Varian) placed in GELoader tips (Eppendorf). The peptides retained were eluted onto a stainless steel target plate with a solution containing 70% acetonitrile, 0.1% trifluoroacetic acid and 10 mg ml^-1 α-cyano-4-hydroxycinnamic acid. The samples were analyzed on an Ultraflex II MALDI-TOF/TOF-mass spectrometer (Bruker Daltonics) operated in the positive reflector mode. For peptide identification, MALDI-TOF spectra were compared to the PilG sequence after in silico digestion with Asp-N and trypsin using the software Biotools v3.0.

Frye S.A., Lång E., Beyene G.T., Balasingham S.V., Homberset H., Rowe A.D., Ambur O.H, & Tønjum T. (2015). The Inner Membrane Protein PilG Interacts with DNA and the Secretin PilQ in Transformation. PLoS ONE, 10(8), e0134954.

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MALDI-TOF/TOF Mass Spectrometry of Tryptic Peptides

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The pooled fractions of peptides were reduced with 10 mM dithiothreitol (DTT) for 1 hour at 60 °C and alkylated with 55 mM iodoacetamide (MP Biomedicals) for 1 hour in the dark at room temperature. Excess iodoacetamide was neutralized with DTT, and then the peptides were digested with trypsin (Promega) at 37 °C for 16 hours. The tryptic digests were desalted with Bond Elut C18 tips and spotted on a MALDI target plate with an equal volume of α-cyano-4-hydroxycinnamic acid (HCCA) matrix (10 mg/mL of 50% ACN containing 0.1% FA). Mass spectra were obtained in reflector positive ion mode using a Bruker Daltonics Ultraflex II MALDI-TOF/TOF mass spectrometer. The MALDI peptide mass fingerprint (PMF) was subjected to tandem MS/MS using MALDI LIFT-TOF/TOF (Bruker Daltonics). Bruker Biotools 3.1 was used to combine PMF and LIFT-MS/MS data and searched with parameters listed below.

Packialakshmi B., Liyanage R., Lay J J.r., Okimoto R, & Rath N. (2015). Prednisolone-Induced Predisposition to Femoral Head Separation and the Accompanying Plasma Protein Changes in Chickens. Biomarker Insights, 10, 1-8.

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