Production of the SFE essential oil used a Speed SFE instrument, model 7070 (Applied Separation Inc. Allentown, PA, USA), consisting of a column oven, air pressure regulator, and a 195 mm × 75 mm i.d. stainless steel column, connected to a NESLAB RTE 7 refrigerated bath (Thermo Electron Corporation, Waltham, MA, USA). Compressed air and CO2 were purchased from Airgas Inc., Radnor, PA, USA. The extraction column was filled with powdered plant material (69 g). Glass wool was added at each end of the column. As modifier, methanol was added at a concentration of 5% to the part of column where the CO2 entered into the column. The extraction temperature was set to 50 °C. Extraction was performed at 250 psi, with a static extract time of 30 min and at a flow rate of 0.5 mL/min, four times for each sample. The SFE extract was then collected in glass vials and stored at −20 °C.
Neslab rte 7
Manufactured by Thermo Fisher Scientific
Sourced in United States
The NESLAB RTE-7 is a recirculating chiller designed for laboratory applications. It provides temperature-controlled fluid circulation for various equipment and processes. The device can maintain a stable temperature within a specified range, as per its technical specifications.
Automatically generated - may contain errors
Lab products found in correlation
F 7000 fluorescence spectrophotometer, by Hitachi (2 mentions)
Fluoressence software, by Horiba (1 mentions)
Prism 7, by GraphPad (1 mentions)
Fluoromax 4 spectrofluorometer, by Horiba (1 mentions)
Synergy htx multi mode microplate reader, by Agilent Technologies (1 mentions)
Bioruptor, by Thermo Fisher Scientific (1 mentions)
Pierce coomassie plus bradford assay kit, by Thermo Fisher Scientific (1 mentions)
M per, by Thermo Fisher Scientific (1 mentions)
Microfuge 22r, by Beckman Coulter (1 mentions)
8 protocols using neslab rte 7
1
Supercritical Fluid Extraction of Essential Oils
2
RNA Thermal Denaturation Kinetics
3
Tryptophan Fluorescence of Nsp12 and Nsp7
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Tryptophan fluorescence spectroscopy was performed using F-7000 Fluorescence Spectrophotometer (Hitachi). The excitation wavelength was set at 280 nm and the emission spectra were recorded from 310 to 370 nm with a 5 nm slit width of excitation and emission. The scan speed was 240 nm/min. The temperature was maintained at 37 °C by a thermostatic water circulator (NESLAB RTE-7; Thermo Scientific). The samples were prepared in 20 mM HEPES, pH 7.5, 65 mM KCl, 5 % glycerol, 2 mM MgCl2, 1 mM DTT. 1 μM Nsp12 and 2 μM Nsp7 was used to record the spectra of Nsp12 and Nsp7, respectively. To record the spectra of Nsp7•12, 1 μM Nsp12 was incubated with 2 μM Nsp7 at 37 °C for 15 min. To collect spectra of denatured proteins, 1 μM Nsp12 was incubated in 8 M urea at room temperature for 1 hour. Three independent measurements, each in three technical replicates, were performed. The same results were obtained with proteins purified three months apart. Means, s.e.m. and second derivatives of the emission spectra were calculated by OriginPro 2021.
4
Fluorescence-based Binding Affinity Assay
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All measurements were conducted on a FluoroMax-4 spectrofluorometer (Horiba Jobin-Yvon). The instrument was equipped with a thermostated cell holder with a Neslab RTE7 water bath (Thermo Scientific). The system was operated by FluorEssence software (version 2.5.3.0 and V3.5, Horiba Jobin-Yvon). All measurements were performed at 20 °C in 25 mM Tris-Cl pH 8, 25 mM NaCl, 1 mM DTT with the exception of measurements in Supplementary Fig. 2f where 25 mM and 100 mM NaCl buffer conditions were compared. CTD peptides were labeled N-terminally with 5,6-carboxyfluorescein (FAM λex = 467 nm, λem = 517 nm; Clonestar) (Supplementary Table 1 ). 10 nM CTD peptide (in a volume of 1.4 ml) was titrated with increasing amounts of SPOC protein. Each data point is an average of three measurements. The binding isotherms were generated by non-liner regression analyses with the software package GraphPad Prism 7 (GraphPad Software, La Jolla).
5
Tryptophan Fluorescence Spectroscopy of Nsp12 and Nsp7
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Tryptophan fluorescence spectroscopy was performed using a model F-7000 fluorescence spectrophotometer (Hitachi). The excitation wavelength was set at 280 nm, and the emission spectra were recorded from 310 to 370 nm, with a 5-nm slit width of excitation and emission. The scan speed was 240 nm/min. The temperature was maintained at 37°C by a thermostatic water circulator (NESLAB RTE-7; Thermo Scientific). The samples were prepared in 20 mM HEPES, pH 7.5, 65 mM KCl, 5% glycerol, 2 mM MgCl2, 1 mM DTT. One micromolar Nsp12 and 2 μM Nsp7 were used to record the spectra of Nsp12 and Nsp7, respectively. To record the spectra of Nsp7·12, 1 μM Nsp12 was incubated with 2 μM Nsp7 at 37°C for 15 min. To collect the spectra of denatured proteins, 1 μM Nsp12 was incubated in 8 M urea at room temperature for 1 h. Three independent measurements, each in three technical replicates, were performed. The same results were obtained with proteins purified 3 months apart. Means, SEM, and second derivatives of the emission spectra were calculated by OriginPro 2021.
6
Whole-Cell Protein Extraction from Thawed Cells
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Extraction of whole-cell protein from thawed cell pellets began by adding 100 μL of cold mammalian protein extraction reagent (M-PER) (Thermo Scientific, ThermoFisher Scientific, Cat#78501) per 10 μL of packed cell volume and 1 μL of 100× halt protease inhibitor cocktail (Thermo Scientific, ThermoFisher Scientific, Cat#87786) per 100 μL of M-PER. After resuspending cell pellets, lysates were sonicated on high for 5 min with 30 s on/off intervals at 4 • C (Diagenode Bioruptor; Thermo Scientific NESLAB RTE-7) to ensure proper lysis of nuclei. Samples were then rotated for 10 min at 4 • C, followed by centrifugation at 14 000g for 15 min at 4 • C (Beckman Coulter Microfuge 22R). Supernatants were transferred to separate tubes and stored at -80 • C for later use. Protein quantification was performed using the Pierce Coomassie Plus Bradford Assay Kit (Thermo Scientific, ThermoFisher Scientific, Cat#23236) with a working range of 125-1500 μg/mL and 96well microplates. Plates were read using a BioTek Synergy HTX Multimode Microplate Reader, which generated a standard curve and protein concentrations for all samples in Microsoft Excel.
7
Anaerobic Digester System Design
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Three identical anaerobic digesters were operated in parallel in this study. Each digester consisted of a 28 L stainless steel conical shape reactor, a peristaltic hose pump (DULCO ® Flex from Prominent Fluid Controls, Australia), a biogas counter (Ritter Company™, MilliGascounter), a thermal probe and a gas trap for biogas sampling. A temperature control unit (Neslab RTE 7, Thermo Fisher Scientific, Newington, USA) was used to maintain the reactor temperature at 35 ± 1 °C. This was achieved by circulating hot water from the temperature control unit through a rubber tube that was firmly wrapped around the reactor.
The reactor and pipeline were encased in polystyrene foam for insulation. The peristaltic hose pump was continuously operated to circulate the digestate at 60 L/h for mixing. Further details of these anaerobic digesters are available elsewhere (Yang et al., 2017) .
The reactor and pipeline were encased in polystyrene foam for insulation. The peristaltic hose pump was continuously operated to circulate the digestate at 60 L/h for mixing. Further details of these anaerobic digesters are available elsewhere (Yang et al., 2017) .
8
Anaerobic Digester Design and Setup
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Three identical anaerobic digesters were used. Each digester (Supplementary Data Figure S1) consists of a 28 L conical shape stainless steel reactor, a peristaltic hose pump (DULCO® Flex from ProMinent Fluid Controls, Australia), a thermal couple with temperature gauge, a custom made gas counter, and a gas trap for biogas sampling. Hot water flowing inside a rubber hose wrapping around the digester was used for heating. The entire reactor was insulated by polystyrene foam. The temperature of the digester was maintained at 35.0±0.5 °C by regulating the temperature inside the rubber hose using a temperature control unit (Neslab RTE 7, Thermo Fisher Scientific, Newington, USA). When necessary, biogas from the gas counter can be directed to a gas trap for biogas composition analysis.
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