Effective binding of a ligand to a protein can be monitored by the ligand induced shift in thermal stability of the protein. Here we present the implementation of a high throughput screening protocol for thermal shift assays in a cellular context, i.e. the ligand induced stabilization is not studied on isolated proteins but instead on endogenous proteins in their normal cellular environment. We refer to the approach as the cellular thermal shift assay (CETSA) and use it for studies of target engagement of drug candidates in a cellular context. The original methodology is based on quantitative detection of remaining soluble intracellular protein after a heat challenge using Western blots (Martinez Molina, Science, 2013). Herein we present and discuss challenges associated with high throughput adaptation of the Western blot protocol and will exemplify this work using human kinases p38a and Erk1/2 and lately also human TS. Whereas the kinase work relies on commercially available detection reagents, the development of a screen compatible CETSA assay for TS first required a careful selection and validation of suitable detection reagents from several mouse and rabbit derived antibodies. The final protocol involves cell treatment, sample handling and detection in low µl volumes in 384-well plates. Results from a screening campaign of a diverse small molecule library of 10.500 compounds on TS will be presented together with confirmation of further validation of identified hit compounds.