Running Protein Thermal Shift Assays With Real-Time PCR

Choosing the wrong buffer doesn't just slow protein characterization down—it can destabilize a protein entirely. Fluorescence-based thermal shift assays enable researchers to screen a full pH range in a single experiment, revealing how protein stability changes across conditions through melt temperature analysis. This application note presents a high-throughput thermal shift assay protocol using a real-time PCR system to map protein stability across a full pH range in a single plate read. The resulting melt temperature data provides a clear, quantitative path to identifying optimal buffer conditions before downstream characterization, formulation, or assay development. Download this application note to explore: Melt curve analysis across 12 pH conditions that identifies the optimal stability window and where destabilization begins How pH and salt concentration influence melt temperature and protein stability How the full protocol configures in real-time PCR software from plate to FRET channel