Protein homeostasis is heavily dependent on a vast network of molecular chaperones that act to prevent the formation and accumulation of misfolded and aggregated forms of proteins. Unfortunately, it is difficult to elucidate the underlying molecular mechanisms of chaperone function using conventional bulk-ensemble techniques, since these tend to mask transient and heterogeneous populations that may be present. Conversely, single-molecule techniques overcome the inherent limitations of ensemble techniques by facilitating the observation of single protein trajectories in real time. Such an approach is therefore ideal for the study of chaperone action. We are developing a protein-folding sensor based on the chloride intracellular channel protein (CLIC) that can be used in single-molecule experiments to report on folding transitions upon interaction with chaperones. To examine the suitability of CLIC1 as a folding-sensor, the unfolding and aggregation of a CLIC1 mutant, CLIC1C98A/C178/C191A/C223A (referred to here as CLIC1CysL), was characterized. We find that CLIC1CysL is more prone to aggregation at elevated temperatures than wild-type CLIC1 as assessed by dynamic light scattering. Furthermore, the small heat shock protein (sHsp) molecular chaperone αB-crystallin (HSPB5) is able to inhibit the heat-induced aggregation of CLIC1CysL. In doing so, αB-crystallin forms a stable, high molecular-mass complex with aggregation-prone CLIC1CysL as assessed by size exclusion chromatography and SDS-PAGE. To develop CLIC1CysL into a protein folding-sensor, strategies that utilize the fluorescence self-quenching of tetramethylrhodamine fluorophores are currently explored. In summary, CLIC1 is a promising candidate as a protein-folding sensor that, with further development, will enable the precise mechanisms of chaperone function to be studied using single-molecule fluorescence techniques.