Autoflex 3 maldi tof tof spectrometer

Manufactured by Bruker

Sourced in Germany, United States

The Autoflex III MALDI-TOF/TOF spectrometer is a mass spectrometry instrument designed for high-performance analysis. It utilizes matrix-assisted laser desorption/ionization (MALDI) and time-of-flight (TOF) technologies to accurately measure the mass-to-charge ratios of analyte molecules.

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Lab products found in correlation

Flexanalysis 3, by Bruker (5 mentions) Protein 1 standard calibration mixture, by Bruker (4 mentions) Groundsteel massive 384 target, by Bruker (3 mentions) Flexcontrol 3, by Bruker (3 mentions) Ziptip c4 micro columns, by Merck Group (3 mentions) Ltq velos, by Thermo Fisher Scientific (1 mentions) Protein calibration standard, by Bruker (1 mentions) Mtp 384 target plate, by Bruker (1 mentions) Zorbax 300sb c18 column, by Agilent Technologies (1 mentions) Fleximaging 2, by Bruker (1 mentions)

Close spelling variants of this product

Autoflex III MALDI-TOF mass spectrometer Autoflex III MALDI-TOF/TOF mass spectrometer Autoflex III Smartbeam MALDI-TOF mass spectrometer Autoflex III smartbeam MALDI-TOF/TOF mass spectrometer Autoflex III MALDI-TOF Autoflex III MALDI-TOF-TOF Autoflex MALDI-TOF Autoflex III MALDI Autoflex III TOF/TOF Autoflex III spectrometer

9 protocols using autoflex 3 maldi tof tof spectrometer

Elucidation of Sulfation and Glycosylation Sites

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Tryptic digestion and detection of the sulfo-and glycopeptides by MS and tandem MS (MS/MS) was performed to elucidate the sulfation and glycosylation sites and the corresponding glycans. The yielded tryptic glycopeptides were purified and enriched for the respective analysis by MALDI-TOF MS where positive ion linear MALDI mass spectra were acquired in different mass ranges and with multiple acquisition conditions on a MALDI-TOF/TOF Autoflex III spectrometer (Bruker Daltonics). Further, the separated glycopeptide-enriched fractions were deglycosylated, and the yielded peptides were also analyzed by MALDI-TOF MS where positive ion reflectron and linear MALDI mass spectra were acquired in different mass ranges and with multiple acquisition conditions on a MALDI-TOF/TOF Autoflex III spectrometer (Bruker Daltonics). This was done for determining which of the theoretical N-glycosylation sites are actually occupied with N-glycans.
For identification of sulfation sites, the obtained deglycosylated peptides were also injected on a nano-HPLC system coupled to an ion trap nano-electrospray mass spectrometer (LTQvelos, ThermoScientific). Acquisition of mass data was performed in a mass range of 300 to 2000 Da excluding monocharged ions. Proteome Discoverer ThermoElectron (v: 1.4) was used to analyze acquired LC-MS/MS data files.

Delignat S., Peyron I., El Ghazaly M., V Kaveri S., Rohde J., Mueller F, & Lacroix-Desmazes S. (2018). Biochemical characterization and immunogenicity of Neureight, a recombinant full-length factor VIII produced by fed-batch process in disposable bioreactors. Cellular immunology, 331.

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MALDI-TOF Protein Desalting and Analysis

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A 10 μl protein sample was desalted using ZipTip^® C4 micro-columns (Merck Millipore, Burlington, MS, USA) and eluted with 0.5 μl SA [sinapinic acid, 10 mg/ml in [70:30] Acetonitrile: Trifluoroacetic acid 0.1%] matrix onto a GroundSteel massive 384 target (Bruker Daltonics, Billerica, MA, USA). An Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics) was used in linear mode with the following settings: 5,000–40,000 Th window, linear positive mode, ion source 1: 20 kV, ion source 2: 18.5 kV, lens: 9 kV, pulsed ion extraction of 120 ns, high gating ion suppression up to 1,000 Mr. Mass calibration was performed externally with protein 1 standard calibration mixture (Bruker Daltonics). Data acquisition, peak peaking and subsequent spectra analysis was performed using FlexControl 3.0 and FlexAnalysis 3.0 software (Bruker Daltonics).

Mikkelsen K., Harwood S.L., Compte M., Merino N., Mølgaard K., Lykkemark S., Alvarez-Mendez A., Blanco F.J, & Álvarez-Vallina L. (2019). Carcinoembryonic Antigen (CEA)-Specific 4-1BB-Costimulation Induced by CEA-Targeted 4-1BB-Agonistic Trimerbodies. Frontiers in Immunology, 10, 1791.

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Pectin Lyase Degradation and MALDI-TOF MS Analysis of AIM Oligosaccharides

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For each AIM sample, 5 mg were suspended in 1 mL of acetate buffer (5 mM, pH 5) and degraded by pectin lyase (0.55 nkatal) prepared according to [59 (link)]. Enzymatic digestion and mass spectrometry acquisition were performed according to [60 (link)]. Briefly, oligosaccharides in the hydrolysates were analysed by MALDI-TOF MS using an Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics, Bremen, Germany). Spectra were recorded in the mass range m/z 600–1400 and exported to Flex Analysis 3.0 software (Bruker) and preprocessed. Ion masses and intensities were normalized according to the ion peak attributed to DU4m4. Oligosaccharide nomenclature was as follows: the letter U corresponds to uronic acid, the following number refers to the number of residues in the oligomer (i.e., DP degree of polymerization), acetyl and methyl esters substitutions were referred to as a and m, respectively, followed by the amount of groups. According to this nomenclature, DU4m4 refers to an oligo-hexouronide of DP4 fully methyl esterified and unsaturated at the nonreducing end. Each AIM sample was analysed in triplicate. Kruskal-Wallis (p-value < 0.05) analyses were performed with R software on MALDI-TOF MS ion intensities.

Segonne S.M., Bruneau M., Celton J.M., Le Gall S., Francin-Allami M., Juchaux M., Laurens F., Orsel M, & Renou J.P. (2014). Multiscale investigation of mealiness in apple: an atypical role for a pectin methylesterase during fruit maturation. BMC Plant Biology, 14, 375.

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MALDI-TOF/TOF Analysis of Protein Aggregation

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MALDI-TOF/TOF studies were performed on a Bruker Autoflex III MALDI-TOF/TOF spectrometer (Bruker, Billerica, MA, USA) equipped with a 200-MHz smart-beam pulsed N2 laser (λ 337 nm). Aliquots of 1 μL of mixtures containing 10 μM or 5 μM protein (Hfra, Yfh1 and/or α-syn), 70 μM AA and 2.5 μM Cu²⁺ or Fe³⁺ prepared in buffer B1, were taken after 0 and 150 min of incubation (25 °C) and supplemented by trifluoroacetic acid (TFA) (0.2% v/v). Samples were then combined with 1 μL of matrix solution (10 μg of sinapinic acid in a solution water:acetonitrile (70:30) containing 0.1% TFA), and a 0.5 μL aliquot of this mixture was spotted onto a steel target plate (MTP 384), air-dried and subjected to mass determination. The IS1 and IS2 voltages were 20 kV and 18.5 kV respectively, and the lens voltage was 7.5 kV. Measurements were performed using a positive reflector mode with matrix suppression below 400 Da. The spectra were calibrated externally using a protein calibration standard (3600–17,000 Da) from Bruker. The experiments were performed in duplicate.

Uceda A.B., Donoso J., Frau J., Vilanova B, & Adrover M. (2021). Frataxins Emerge as New Players of the Intracellular Antioxidant Machinery. Antioxidants, 10(2), 315.

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MALDI-TOF/TOF Analysis of Protein Samples

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A 2 μl protein sample was desalted using ZipTip® C4 micro-columns (Merck Millipore) and eluted with 0.5 μl sinapinic acid (10 mg/ml in [70:30] Acetonitrile: Trifluoroacetic acid 0.1%) matrix onto a GroundSteel massive 384 target (Bruker Daltonics). An Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics) was used in linear mode with the following settings: 5000–40,000 Th window, linear positive mode, ion source 1: 20 kV, ion source 2: 18.5 kV, lens: 9 kV, pulsed ion extraction of 120 ns, high gating ion suppression up to 1000 Mr. Mass calibration was performed externally with protein 1 standard calibration mixture (Bruker Daltonics). Data acquisition, peak peaking, and subsequent spectra analysis was performed using the FlexControl 3.0 and FlexAnalysis 3.0 software (Bruker Daltonics).

Compte M., Harwood S.L., Muñoz I.G., Navarro R., Zonca M., Perez-Chacon G., Erce-Llamazares A., Merino N., Tapia-Galisteo A., Cuesta A.M., Mikkelsen K., Caleiras E., Nuñez-Prado N., Aznar M.A., Lykkemark S., Martínez-Torrecuadrada J., Melero I., Blanco F.J., Bernardino de la Serna J., Zapata J.M., Sanz L, & Alvarez-Vallina L. (2018). A tumor-targeted trimeric 4-1BB-agonistic antibody induces potent anti-tumor immunity without systemic toxicity. Nature Communications, 9, 4809.

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MALDI-TOF Analysis of Sap1 Fragment

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The stable fragment of Sap1 obtained following thermolysin treatment was purified by reverse phase HPLC on a ZORBAX 300SB C18 column (Agilent). The molecular mass of the Sap1 stable fragment was determined by MALDI-TOF mass spectrometry. For MALDI-TOF the sample was spotted using sinapinic acid [10 mg/ml] in [70:30] acetonitrile-trifluoroacetic acid [0.1%]) on a MTP 384 target plate (Bruker Daltonics). An Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics) was used in linear mode with the following settings: 5,000- to 50,000-Da window; linear positive mode; ion source 1, 20 kV; ion source 2, 18.5 kV; lens, 9 kV; pulsed ion extraction of 120 ns; and high gating ion suppression up to 1,000 M_r. Mass calibration was performed externally with Bruker’s Protein 1 standard calibration mixture (Bruker Daltonics). Data acquisition was performed using the FlexControl 3.0 software program (Bruker Daltonics), and peak searching and subsequent spectral analysis were performed using FlexAnalysis 3.0 software (Bruker Daltonics).

Jørgensen M.M., Ekundayo B., Zaratiegui M., Skriver K., Thon G, & Schalch T. (2018). Structure of the replication regulator Sap1 reveals functionally important interfaces. Scientific Reports, 8, 10930.

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MALDI-TOF Protein Desalting and Analysis

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A 2 μl protein sample was desalted using ZipTip® C4 micro-columns (Millipore) and eluted with 0.5 μl SA (sinapinic acid, 10 mg/ml in [70:30] Acetonitrile: Trifluoroacetic acid 0.1%) matrix onto a GroundSteel massive 384 target (Bruker Daltonics, Billerica, MA, USA). An Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics) was used in linear mode with the following settings: 5000–40000 Th window, linear positive mode, ion source 1: 20 kV, ion source 2: 18.5 kV, lens: 9 kV, pulsed ion extraction of 120 ns, high gating ion suppression up to 1000 Mr. Mass calibration was performed externally with protein 1 standard calibration mixture (Bruker Daltonics). Data acquisition, peak peaking and subsequent spectra analysis was performed using FlexControl 3.0 and FlexAnalysis 3.0 software (Bruker Daltonics).

Alvarez-Cienfuegos A., Nuñez-Prado N., Compte M., Cuesta A.M., Blanco-Toribio A., Harwood S.L., Villate M., Merino N., Bonet J., Navarro R., Muñoz-Briones C., Sørensen K.M., Mølgaard K., Oliva B., Sanz L., Blanco F.J, & Alvarez-Vallina L. (2016). Intramolecular trimerization, a novel strategy for making multispecific antibodies with controlled orientation of the antigen binding domains. Scientific Reports, 6, 28643.

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MALDI-TOF Imaging of Wheat Sections

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All MSI measurements were performed in an Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonik) equipped with a Smartbeam laser (355nm, 200 Hz) and controlled using the Flex Control 3.0 software package. The mass spectrometer was operated with positive polarity in reflectron mode, and spectra were acquired in the range of 500–2000 m/z.
The laser raster size was set at 100 μm, which is approximately equal to the laser spot diameter. At this resolution, it takes approximately 30min to complete an image of one wheat section. The signal was initially optimized by manually adjusting the laser power and the number of laser shots fired. According to this procedure, full-scan MS experiments were run with 200 laser shots per step and using the laser power that generated the best signal-to-noise ratio. Image acquisition at tissue surfaces was performed using Flex Imaging 2.1 software (Bruker Daltonik). Relative comparisons of the released oligosaccharides in the different tissue sections and of the ratios of different ions in the same tissue section were performed through labelled normalization using MALDI Tools 1.1 software (Källback et al., 2012 (link)) compatible with Flex Imaging 2.1.

Veličković D., Ropartz D., Guillon F., Saulnier L, & Rogniaux H. (2014). New insights into the structural and spatial variability of cell-wall polysaccharides during wheat grain development, as revealed through MALDI mass spectrometry imaging. Journal of Experimental Botany, 65(8), 2079-2091.

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PsaA Antigen Functionalization and Characterization

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For functionalization of PsaA, 1.35 mg of SATA was incorporated at 45 minute-intervals for 3 times (0.45 mg x 3, ~40 equiv) into 5 mg/mL of PsaA antigen in PBS 0.1 M pH 7.4, at 4ºC under stirring. Subsequently, the reaction mixture was dialyzed in 0.1 M PBS to eliminate the excess of reagent using dialysis membrane (3.5 kDa) at 4°C. The ability of the conjugated protein to retain its structural conformation was studied using circular dichroism (CD) using Jasco J-810 spectropolarimeter. The baseline was obtained using a blank solution containing the buffer. Solution of PsaA or the conjugated PsaA-SATA was diluted to 0.1 mg/mL in buffer solution and was scanned over the wavelength range 200-260 nm. The analysis was performed using a 1-nm quartz cylindrical cell at a scanning speed of 50 nm/min, 0.1 nm bandwidth and a temperature of 20°C. PsaA and PsaA-SATA mass spectra were also determined on an Autoflex III MALDI-TOF/TOF spectrometer (Bruker Daltonics).

Robla S., Prasanna M., Varela-Calviño R., Grandjean C, & Csaba N. (2021). A chitosan-based nanosystem as pneumococcal vaccine delivery platform. Drug delivery and translational research, 11(2).

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