Far-UV (250–200 nm) CD protein spectra were recorded at 20°C and 95°C using a JASCO J-815 spectropolarimeter, in a 1 mm optical path length cuvette. Denaturation spectra were performed at 220 nm, heating from 20 to 95°C with an increase of 1°C min−1. Protein samples (40 µM unless otherwise indicated) were prepared in 200 µl of 50 mM potassium phosphate buffer (pH 8, unless otherwise indicated). The temperature was regulated using a Peltier-type JASCO CDF-426S/15 thermostatic controller. All experiments were performed in triplicate. The data were corrected from the background and extracted using the Spectra Manager Suite (JASCO). Mean values and s.e. of the CD spectra were analysed and plotted using Graph Pad Prism 7.0. The denaturation graphs present all data points from the triplicate measurements.
Spectra manager suite
Manufactured by Jasco
Sourced in United States
Spectra Manager Suite is a software application that provides a comprehensive platform for analyzing and managing spectral data. It offers a range of tools and features to facilitate the processing, visualization, and interpretation of spectral information.
Automatically generated - may contain errors
Lab products found in correlation
4 protocols using spectra manager suite
1
Far-UV Circular Dichroism of Protein Denaturation
2
Circular Dichroism Analysis of Exenatide
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Circular dichroism was used to determine exenatide’s secondary structure. Exenatide samples were analyzed at a concentration of 0.125 mg/mL on a J-815 (JASCO, Easton, MD, USA). Instrument sample temperature was held at 37 °C using a Peltier attachment. Circular dichroism spectra were averaged from 5 collected scans over a wavelength range of 245 to 195 nm, with an accumulation rate of 20 nm/min and a data integration time (DIT) of 4 s. Percent contributions from α-helix, β-sheet and unordered secondary structures were quantified using a third-party Spectra Manager Suite (JASCO) add-in, CDPro analysis. In this software, the CONTIN method was chosen, using soluble-membrane protein 56 (SMP 56) as the reference protein for secondary structural analysis.
3
Far-UV CD Protein Spectra Analysis
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Far-UV CD protein spectra (250–200 nm) were recorded at 25 °C using a JASCO J-815 spectropolarimeter in a 1-mm optical path length cuvette. Fifteen-micromolar protein samples were dissolved in 20 mM phosphate buffer (pH 7.4) containing 0.3 M NaF in a total volume of 200 μL. The temperature was maintained by a Peltier-type JASCO CDF-426S/15 thermostatic controller. Raw spectral data were extracted using the Spectra Manager Suite (JASCO). All experiments were performed in triplicate. Mean values and standard error of the mean (SEM) for each normalized data point were plotted and analyzed using GraphPad Prism 9.0. To account for small experimental variations in protein concentration during sample preparation, molar ellipticity (θ, mdeg) was normalized using the following equation:
4
Determination of Total Phenol Content
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The total phenol content (TPC) was determined through Folin–Ciocalteau’s method, using gallic acid as standard (Sigma-Aldrich, St. Louis, MO, USA). Briefly, 0.125 mL of sample (properly diluted with water to obtain an absorbance value within the linear range of the spectrophotometer) underwent an addition of: 0.5 mL of distilled water, 0.125 mL of Folin–Ciocalteau’s (Sigma-Aldrich, St. Louis, MO, USA) reagent and 1.25 mL of an aqueous solution of Na2CO3 7.5% (w/v%), bringing the final volume to 3 mL with water. After mixing, the samples were kept in the dark for 90 min. After the reaction period, the absorbance was measured at 760 nm using a V-730 UV-visible/NIR spectrophotometer operated by Spectra Manager™ Suite (Jasco Inc., Easton, MD, USA). Each sample was analyzed in triplicate, and the concentration of total polyphenols was calculated in terms of gallic acid equivalents (GAE) [14 (link)].
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