eBooks & Guides

Characterizing protein stability, powerful approaches to study protein denaturation

Issue link: https://resources.nanotempertech.com/i/1050533

Contents of this Issue

Navigation

Page 11 of 24

9 How it works In contrast to conventional DSF methods in which fluorescence labeling is required, nanoDSF measures intrinsic tryptophan or tyrosin fluorescence to monitor protein unfolding. Both the fluorescence intensity and the fluorescence maximum strongly depends on the close surroundings of the tryptophan. Therefore, the ratio of the fluorescence intensities at 350 nm and 330 nm is suitable to detect any changes in protein structure, for example due to protein unfolding. It can be used to characterize thermal stability or unfolding, chemical denaturation, and relative aggregation. Strengths With this technique, protein stability can be monitored under native conditions. Also, no sample prep is needed and little sample is consumed. Another beneficial feature of the technology is the robust data density or the number of readings collected during each data point that translates into higher quality and more reliable information collected during each experiment compared to other methodologies. Weaknesses There are trade-offs when considering nanoDSF vs other methodologies. There are methods that provide higher throughput (DSF/Thermofluor) but require labeling of samples and don't provide highly quantitative results. There are systems available that can perform multiple analysis on protein stability, however, the precision of each method may be compromised vs. nanoDSF. Researchers need to consider the quality and amount of data generated and required when evaluating the weaknesses of nanoDSF. Conclusion NanoDSF is a label-free approach which uses rapid sample scanning and intrinsic fluorescence detection to collect highly precise and accurate thermal unfolding profiles in a high-throughput manner. Nano differential scanning fluorimetry (nanoDSF) is a modified version of DSF that measures protein stability using intrinsic tryptophan or tyrosin fluorescence. nanoDSF

Articles in this issue

view archives of eBooks & Guides - Characterizing protein stability, powerful approaches to study protein denaturation