# Differential Scanning Fluorimetry (DSF) ## expand\_more [**CMI DSF resources and user guides**](/node/750981#dsf-resources)**⬇︎** Differential Scanning Fluorimetry (DSF) measures protein unfolding by monitory changes in fluorescence as a function of temperature. **Conventional DSF** uses a hydrophobic fluorescent dye that binds to proteins as they unfold. **NanoDSF** measures changes in intrinsic protein fluorescence as proteins unfold. ![Diagram of Differential Scanning Fluorimetry](/sites/g/files/omnuum4016/files/2026-04/DSF%20diagram%202026.png) ### Conventional DSF Conventional **Differential Scanning Fluorimetry (DSF)** uses a real-time PCR instrument to monitor thermally induced protein denaturation by measuring changes in fluorescence of a dye that binds preferentially to unfolded protein (such as Sypro Orange, which binds to hydrophobic regions of proteins exposed by unfolding). This experiment is also known as a [Protein Thermal Shift Assay](http://www.lifetechnologies.com/us/en/home/life-science/pcr/real-time-pcr/real-time-pcr-applications/real-time-pcr-protein-analysis/protein-thermal-shift.html?icid=fr-proteinmelt), because shifts in the apparent melting temperature can be measured upon the addition of stabilizing or destabilizing binding partners or buffer components. ![QuantStudio 6](/sites/g/files/omnuum4016/files/styles/hwp_1_1__100x100_scale/public/2025-02/QuantStudio6_image_0.png?itok=kOMn_ZBL) The CMI has a modified [Life Technologies Quant Studio 6/7](https://www.thermofisher.com/us/en/home/life-science/pcr/real-time-pcr/real-time-pcr-instruments/quantstudio-6-flex-real-time-pcr-system.html), for conventional DSF. The CMI has the Life Technologies Protein Thermal Shift Analysis software for DSF data fitting. ### NanoDSF **NanoDSF** is a modified differential scanning fluorimetry method which monitors intrinsic tryptophan and tyrosine fluorescence as a function of temperature, time, or denaturant concentration. Tryptophan and tyrosine fluorescence intensity and wavelength maximum will vary as the local chemical environment changes, with significant changes occurring as buried or packed aromatic side chains become solvent exposed upon unfolding. NanoDSF measures fluorescence intensity at 350 nm and 330 nm and compares the ratio as a function of temperature or denaturant concentration. NanoDSF can be used for a broader range of protein samples than traditional DSF and has significantly higher throughput and lower sample consumption than DSC or CD. Free energies of folding and temperatures of unfolding measured using NanoDSF are comparable to values determined by DSC for a range of sample types. The only significant limitation is that the protein of interest must contain aromatic amino acids (tryptophan or tyrosine). ![Prometheus NT.plex](/sites/g/files/omnuum4016/files/styles/hwp_1_1__100x100_scale/public/2025-02/prometheus-nt.plex_.png?itok=_Q9-ZXeC) The CMI has a **Prometheus NT.Plex instrument** from [NanoTemper Technologies](https://nanotempertech.com/) with aggregation optics. The CMI has these Data collection and analysis software packages: PR.ThermControl for thermal stability data collection, PR.ChemControl for chemical stability data collection, PR.TimeControl for time interval data collection, and PR.Stability Analysis for advanced data analysis. Conventional DSF Resources Quant Studio DSF Supplies QPCR Resources ## Conventional DSF Resources [CMI QuantStudio DSF Getting Started Guide](/sites/g/files/omnuum4016/files/2026-01/CMI%20QuantStudio%20DSF%20Getting%20Started%20Guide.pdf "CMI QuantStudio DSF Getting Started Guide") [Protein Thermal Shift Studies manual](/sites/g/files/omnuum4016/files/cmi/files/abi_protein_thermal_shift_manual.pdf "ABI protein thermal shift manual") from Applied Biosystems by Life Technologies ## Quant Studio DSF Supplies All Experiments: - 96-well FAST-block optical plate, eg.: LifeTechnologies MicroAmp FAST optical 96-well reaction plate, 0.1 ml, 4346907 - optical adhesive film, eg.: LifeTechnologies MicroAmp Optical Adhesive Film, 4360954 DSF/Protein Thermal Shift Experiments - DSF compatible dye, eg.: LifeTechnologies Protein Thermal Shift Dye Kit, 4461146 (Sypro Orange) - samples, ligands, buffers qPCR Experiments - qPCR reagents (eg. LifeTechnologies PowerUp SYBR Green Master Mix, A25742) - primers and templates ## QPCR Resources [CMI QuantStudio qPCR Getting Started Guide](/sites/g/files/omnuum4016/files/2025-03/CMI%20QuantStudio%20qPCR%20Getting%20Started%20Guide.pdf "CMI QuantStudio qPCR Getting Started Guide") [Quant Studio 6/7 Quick Reference Guide](http://tools.thermofisher.com/content/sfs/manuals/4489826.pdf) from Applied Biosystems by Life Technologies NanoDSF Resources Prometheus NT.Plex Supplies ## NanoDSF Resources [CMI Prometheus NanoDSF Getting Started Guide](/sites/g/files/omnuum4016/files/2025-03/CMI%20Prometheus%20NanoDSF%20Getting%20Started%20Guide.pdf "CMI Prometheus NanoDSF Getting Started Guide") [CMI Guide to Prothetheus NT.Plex Capillaries](/sites/g/files/omnuum4016/files/cmi/files/cmi_guide_to_prometheus_ntplex_capillaries.pdf "CMI Guide to Prometheus NTPlex Capillaries") [NanoDSF Technology Page](https://nanotempertech.com/prometheus/) NanoTemper Technologies ## Prometheus NT.Plex Supplies - NT.Plex Capillary Chips - 2x 8 Standard 24-Capillary Chips, NanoTemper Catalog # PR-AC002 - 2x 8 High Sensitivity 24-Capillary Chips, NanoTemper Catalog # PR-AC006 - 384-well plates for loading capillaries - Protein samples, ligands, buffers [ Data Management and File Formats arrow\_circle\_right ](/data-management) [ DSF Services arrow\_circle\_right ](/dsf-services) [ About Data Collection Services arrow\_circle\_right ](/services) See also:- [ molecular properties ](/cmi-terms/molecular-properties) - [ Differential Scanning Fluorimetry ](/cmi-terms/differential-scanning-fluorimetry)