All NMR experiments were performed on a Bruker Bruker Avance III 700 MHz spectrometer, except for the 3D assignment of β2AR-Cter performed on a 800 MHz, and for the 3JHNHA of GHSR-Cter performed on a 500 MHz. The 700 MHz and 800 MHz spectrometers are equipped with a cryogenic triple-resonance (1H, 15N, 13C) probe and shielded z-gradients. All NMR experiments were recorded at 20 °C in a buffer (named NMR buffer) composed of 50 mM Bis-Tris pH 6.7, 150 mM NaCl, 1 mM EDTA, 0.5 mM TCEP, 5% D2O (Eurisotop), and 5 mM DSS-d6 (2,2-dimethyl-2-silapentane-5-sulfonate, Sigma) as internal reference [42 ]. All experiments used the pulse sequences provided by Bruker TOPSPIN 3.2. Squared cosine apodization was used in indirect dimensions, prior to zero-filling and Fourier transformation using TOPSPIN (version 4.0.6, Bruker) and data processing was performed using NMRFAM-SPARKY (version 1.414, [43 (link)]). For each NMR experiments, concentrations of GPCR-Cters were indicated in Table S1 . For all NMR experiments, data were measured for all residues of C-terminus regions excepted proline residues, the residue A339 of β2AR-Cter, and the first N-terminal residue.
Dss d6
Manufactured by Merck Group
Sourced in United States
The DSS-d6 is a laboratory equipment product manufactured by Merck Group. It serves as a core analytical tool for various scientific applications. The DSS-d6 functions as a deuterium-enriched solvent for nuclear magnetic resonance (NMR) spectroscopy analysis.
Automatically generated - may contain errors
Lab products found in correlation
Nmrfam sparky, by Bruker (1 mentions)
Topspin, by Bruker (1 mentions)
Topspin 3, by Bruker (1 mentions)
U 13c6 glucose, by Cambridge Isotopes (1 mentions)
Doxycycline dox, by Merck Group (1 mentions)
5 13c glutamine, by Cambridge Isotopes (1 mentions)
Sodium acetate d3, by Merck Group (1 mentions)
Acetic acid d4, by Merck Group (1 mentions)
Accq tag amino acid analysis method, by Waters Corporation (1 mentions)
Avance spectrometer, by Bruker (1 mentions)
Nmr suite 8, by Chenomx (1 mentions)
4 protocols using dss d6
1
NMR Spectroscopy of GPCR C-Termini
2
Isotopic Labeling for Metabolomic Analysis
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Chemicals were used without further purification form commercial vendors: 5-13C-glutamine (Cambridge Isotopes), U-13C6-glucose (Cambridge Isotopes), Doxycycline (Dox, Sigma), D2O (Sigma) and DSS-d6 (3-(trimethylsilyl)-1-propanesulfonic acid-d6 disodium salt, Sigma).
3
Scotch Whisky Provenance Analysis
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Scotch Whisky samples (n=148) were provided by the Scotch Whisky Research Institute (SWRI). Every two years the SWRI requests around 50 Scotch Whisky brands from its member companies. These brands are selected by the SWRI to represent the breadth and volume of Scotch Whisky in production and are sourced directly from the producers to guarantee provenance. The Scotch Whisky samples in this work were subsampled (15 ml in sealed vials) at the SWRI, from the carefully maintained sample sets collected in 2010, 2012 and 2014, and transferred to University of Edinburgh. These sampled brands represented (r, %) of Scotch Whisky UK and export case sales (2010, n=47, r=67 %; 2012, n=49, r=71 %; and 2014, n=52, r=73%) . For a complete list of anonymised samples see Table S1 in Supplementary Material. Known counterfeit samples (n=32) were also provided by the SWRI. Their origin and counterfeit status determination is discussed in the Supplementary Material. DSS-d6 (98 %), D2O (99 %), acetic acid-d4 (99.5 %), and sodium acetate-d3 (99 %) were acquired from Sigma-Aldrich Co.
4
Quantification of Metabolites in Cell Culture
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Glucose and lactate concentrations in culture supernatants were measured with an automated YSI 7100 Multiparameter Bioanalytical System (Dayton, OH, USA). Amino acid concentrations were analyzed by HPLC using the Waters AccQ.Tag Amino Acid Analysis Method (Waters, Milford, MA) as described elsewhere [9] . The extracellular concentrations of citrate and pyruvate were determined by 1 H-NMR spectroscopy using a 500 MHz Avance spectrometer (Bruker, Billerica, MA) with a 5 mm QXI inversed probe. Spectra were recorded at 25 °C using a NOESY-based pulse sequence with water pre-saturation, performing 256 scans with 4 s acquisition time, 2 s relaxation delay and 100 ms mixing time. DSS-d6 (Sigma Aldrich, St. Louis, MO; USA) was used as internal standard for metabolite quantification in all samples. In order to obtain a similar pH between samples, they were mixed with phosphate buffer (pH 7.4) prepared in D 2 O at a 2:1 ratio. Before spectra acquisition, the spectrometer was calibrated by determining the 908 pulse and the water chemical shift centre of each sample. Each spectrum was phased, baseline corrected and integrated using the Chenomx NMR Suite 8.0 (Chenomx Inc., Edmonton, Alberta, Canada) software.
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