Esp 300 x band spectrometer

Manufactured by Bruker

Sourced in Germany

The ESP-300 X-band spectrometer is a laboratory instrument designed for electron spin resonance (ESR) spectroscopy. It operates in the X-band frequency range and is used for the detection and analysis of paramagnetic species in materials. The core function of the ESP-300 is to measure the absorption of microwave radiation by a sample placed in a strong magnetic field, providing information about the electronic structure and properties of the sample.

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7 protocols using esp 300 x band spectrometer

EPR Analysis of Gamma-Irradiated Explants

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Electron paramagnetic resonance (EPR) spectroscopy visualizes the EPR-active centers in the γ-irradiated samples. In this study it was applied to characterize EPR spectra with particular attention to the bone-characteristic EPR-signals in the explants harvested from animals. In order to generate active centers in the investigated samples, explants were exposed to the gamma irradiation using ⁶⁰Co gamma source Issledovatiel with dose rate of 1 kGy/h and a total dose of 35 kGy. To distinguish the signals coming from the scaffolds and from the tissues, EPR spectra coming from the non-implanted scaffolds after radiation sterilization were measured as well.
EPR analysis was performed using a Bruker ESP300 X-band spectrometer equipped with high-sensitivity cavity ER 4108 TMH, Bruker ER035 M Gaussmeter and microwave frequency meter HP 5342A. For absolute g-value determination, a calibration using diphenylpicrylhydrazyl (DPPH) at 0.1 mW (g = 2.0036) was executed. The analysis was performed at the room temperature, with modulation frequency of 100 kHz, modulation amplitude of 0.1 mT and microwave power of 10 mW.

Chróścicka A., Jaegermann Z., Wychowański P., Ratajska A., Sadło J., Hoser G., Michałowski S, & Lewandowska-Szumiel M. (2015). Synthetic Calcite as a Scaffold for Osteoinductive Bone Substitutes. Annals of Biomedical Engineering, 44, 2145-2157.

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X-band EPR Spectroscopy at 77K

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Continuous wave X-band EPR
spectra were recorded at 77 K in the MIT Department of Chemistry Instrumentation
Facility on a Bruker ESP-300 X-band spectrometer equipped with a quartz
finger Dewar filled with liquid N₂. Experimental conditions
were as follows: microwave frequency, 9.45 GHz; modulation amplitude,
0.15 mT; modulation frequency, 100 kHz; time constant, 5.12 ms; scan
time, 41.9 s; microwave power, 20 μW.

Wei Y., Li B., Prakash D., Ferry J.G., Elliott S.J, & Stubbe J. (2015). A Ferredoxin Disulfide Reductase Delivers Electrons to the Methanosarcina barkeri Class III Ribonucleotide Reductase. Biochemistry, 54(47), 7019-7028.

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Hydroxyl Radical Detection in Cyanobacteria

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Spin trapping assays with the spin probe 4-pyridyl-1-oxide-N-tert-butylnitrone (4-POBN; Sigma-Aldrich, St. Louis, USA) to detect the formation of hydroxyl radicals^{31 (link)} were carried out using cyanobacterial cells (OD₇₃₀ = 0.65) in 10 mM sodium phosphate buffer (pH 7.0) containing 50 mM 4-POBN, 4% (v/v) ethanol, 50 µM Fe-EDTA, and 50 µM MV. After a 30-min incubation in the light (10 µmol m⁻² s⁻¹), the samples were centrifuged at 7000g for 1 min, and the supernatants were frozen in liquid nitrogen and stored at − 80 °C for electron paramagnetic resonance (EPR) spectra analysis. The EPR spectra were recorded at room temperature in a standard quartz flat cell using an ESP-300 X-band spectrometer (Bruker, Rheinstetten, Germany). The following parameters were used: microwave frequency, 9.73 GHz; modulation frequency, 100 kHz; modulation amplitude, 1 G; microwave power, 6.3 milliwatt; receiver gain, 2 × 104; time constant, 40.96 ms; number of scans: 4.

Tanaka K., Shimakawa G, & Nakanishi S. (2020). Time-of-day-dependent responses of cyanobacterial cellular viability against oxidative stress. Scientific Reports, 10, 20029.

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Continuous Wave EPR Spectroscopy at 77K

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Continuous wave (cw) X-band
EPR spectra were recorded at 77 K in the MIT Department of Chemistry
Instrumentation Facility on a Bruker ESP-300 X-band spectrometer equipped
with a quartz finger Dewar filled with liquid N₂. Experimental
conditions were as follows: microwave frequency, 9.45 GHz; modulation
amplitude, 0.15 mT; modulation frequency, 100 kHz; time constant,
5.12 ms; and scan time, 41.9 s. A microwave power of 10 μW and
an average of 10 scans was used for G•, while a power of 160
μW and 100 scans was used for the thiosulfurnanyl radical.

Wei Y., Mathies G., Yokoyama K., Chen J., Griffin R.G, & Stubbe J. (2014). A Chemically Competent Thiosulfuranyl Radical on the Escherichia coli Class III Ribonucleotide Reductase. Journal of the American Chemical Society, 136(25), 9001-9013.

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Spin-Trapping Analysis of Reactive Oxygen Species

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Spin-trapping assays with TEMPD-HCl (Sigma-Aldrich, St. Louis, USA) to detect 1 O 2 (Krieger-Liszkay et al., 2015; Ramel et al., 2013a) or the spin probe 2diethylphosphono-2-methyl-3,4-dihydro-2H-pyrrole 1-oxide (DEPMPO) to detect the formation of O 2 •-and HO • (Frejaville et al., 1995; Heyno et al., 2009) were carried out using thylakoid membranes at a concentration of 10 μg Chl mL -1 . The thylakoids were suspended in a buffer containing 0.3 M sorbitol, 50 mM KCl, 5 mM MgCl 2 , 25 mM HEPES pH 7.6. Samples were illuminated for 2 min with red light (RG 630) (1000 μE m -2 s -1 ) in the presence of 100 mM TEMPD-HCl or in the presence of 50 mM DEPMPO. In the DEPMPO assay 50 μM diethylenetriaminepentaacetic acid was added and the thylakoids were uncoupled. EPR spectra were recorded at room temperature in a standard quartz flat cell using an ESP-300 X-band spectrometer (Bruker, Rheinstetten, Germany). The following parameters were used: microwave frequency, 9.73 GHz; modulation frequency, 100 kHz; modulation amplitude, 1 G; microwave power, 63 mW in TEMPD assays; receiver gain, 2×10 4 ; time constant, 40.96 ms and number of scans 4.
The background EPR signals of TEMPD and DEPMPO were subtracted using the same experimental conditions without adding thylakoids. For control experiments, D 2 O and NaN 3 were used to enhance or to quench 1 O 2 production and superoxide dismutase

Sánchez-Corrionero Á., Sánchez-Vicente I., González-Pérez S., Corrales A., Krieger-Liszkay A., Lorenzo Ó, & Arellano J.B. (2017). Singlet oxygen triggers chloroplast rupture and cell death in the zeaxanthin epoxidase defective mutant aba1 of Arabidopsis thaliana under high light stress. Journal of plant physiology, 216.

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EPR Spectroscopy of Nitrogenase Proteins

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EPR spectra were recorded using a Bruker X-band ESP 300 spectrometer equipped with a Bruker dual-mode cavity (ER4116DM) and an Oxford Instruments ESR 900 continuous flow cryostat. Measurements utilized a 2 mW microwave power and 0.746 mT/100 kHz modulation at temperatures of 10 K. Spectra were analyzed using the software package EasySpin (version 5.1.9) as implemented in Matlab [39 (link)]. MoFe^rec and FeP^rec were prepared by reconstitution of the lyophilized proteins into a buffer solution containing 20 mM Tris, 100 mM NaCl, and 2.5 mM dithionite (pH 7.3). MoFe^lyo and FeP^lyo were transferred into X-band EPR tubes manually using a custom-fitted funnel. All EPR sample preparations were performed under an anoxic N₂ atmosphere.

Van Stappen C., Decamps L, & DeBeer S. (2020). Preparation and spectroscopic characterization of lyophilized Mo nitrogenase. Journal of Biological Inorganic Chemistry, 26(1), 81-91.

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Anaerobic EPR Spectroscopy of Reduced Proteins

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EPR spectra were recorded with an X-band ESP 300 spectrometer (Bruker). Samples were loaded in quartz EPR tubes (707SQ, 250 mm) under strictly anaerobic conditions and frozen in liquid nitrogen. To obtain reduced protein, 10 mM sodium dithionite was added under strict anaerobic condition. The EPR spectra were recorded at 10 K, with a microwave power of 28 dB and 0.47 mW at highest signal intensity. The shown spectra are an average of 16 scans each, with a total scan time of 22 min per spectrum.

Wang C., Nehls C., Baabe D., Burghaus O., Hurwitz R., Gutsmann T., Bröring M, & Kolbe M. (2021). Flagellin lysine methyltransferase FliB catalyzes a [4Fe-4S] mediated methyl transfer reaction. PLoS Pathogens, 17(11), e1010052.

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