This review discusses various methods for monitoring protein stability, with a focus on fluorescence-based techniques. Proteins are essential biomolecules whose structure determines function, and their stability is influenced by environmental factors. Many label-free methods exist for studying protein stability, but fluorescence-based techniques are particularly favored for their low instrument and expertise requirements. Thermal shift assays (TSAs), also known as differential scanning fluorimetry (DSF) or ThermoFluor, are highlighted as a practical and accessible method for assessing protein stability. These assays involve monitoring fluorescence changes as temperature increases, with the melting temperature (Tm) indicating the point at which a protein begins to unfold. TSA is compared to isothermal chemical denaturation (ICD), and both methods are discussed in terms of their advantages and challenges. The review also explores other techniques such as circular dichroism (CD), nuclear magnetic resonance (NMR), and differential scanning calorimetry (DSC), which are more equipment-intensive but provide detailed thermodynamic data. Fluorescence-based methods like DSF are noted for their high throughput and suitability for screening large numbers of compounds, making them valuable in pharmaceutical research. The review emphasizes the importance of optimizing buffer conditions and minimizing interferences to ensure accurate results. It also discusses the use of intrinsic fluorescence and external dyes, as well as the advantages of label-free methods in studying protein-ligand interactions and stability. Overall, the review highlights the importance of selecting the appropriate method for protein stability analysis, considering factors such as throughput, sensitivity, and equipment availability.This review discusses various methods for monitoring protein stability, with a focus on fluorescence-based techniques. Proteins are essential biomolecules whose structure determines function, and their stability is influenced by environmental factors. Many label-free methods exist for studying protein stability, but fluorescence-based techniques are particularly favored for their low instrument and expertise requirements. Thermal shift assays (TSAs), also known as differential scanning fluorimetry (DSF) or ThermoFluor, are highlighted as a practical and accessible method for assessing protein stability. These assays involve monitoring fluorescence changes as temperature increases, with the melting temperature (Tm) indicating the point at which a protein begins to unfold. TSA is compared to isothermal chemical denaturation (ICD), and both methods are discussed in terms of their advantages and challenges. The review also explores other techniques such as circular dichroism (CD), nuclear magnetic resonance (NMR), and differential scanning calorimetry (DSC), which are more equipment-intensive but provide detailed thermodynamic data. Fluorescence-based methods like DSF are noted for their high throughput and suitability for screening large numbers of compounds, making them valuable in pharmaceutical research. The review emphasizes the importance of optimizing buffer conditions and minimizing interferences to ensure accurate results. It also discusses the use of intrinsic fluorescence and external dyes, as well as the advantages of label-free methods in studying protein-ligand interactions and stability. Overall, the review highlights the importance of selecting the appropriate method for protein stability analysis, considering factors such as throughput, sensitivity, and equipment availability.