Real-time detection of T cell activation by visualizing TCR nanoclusters with a cholesterol derived...

Successful T-cell based immunotherapy usually depends on the activation of T cells. Most of commonly used methods for assessing T cell activity rely on the antibody-based technology, which focus on detecting protein-centered activation markers, including CD25, cytokines and so on. However, these methods always involve tedious sample-preparation process, labor-consuming and costly, which could not be utilized in real-time detection. The T cell receptor (TCR) clustering is another kind of essential T cell activation marker on the membrane, which increases during the activation state of T cells. We herein developed a cholesterol derived aggregation-induced emission (AIE) fluorescent probe (R-TPE-PEG-Chol) for detecting T cell activation in real-time. Five probes were first designed and synthesized and among them COOH-TPE-PEG-Chol displayed the best imaging effects, which had no significant impact on the key physiological functions of T cells. In addition, we have proved that COOH-TPE-PEG-Chol was introduced onto the naïve T cell membrane in its molecularly dissolved form without fluorescent emission. While during T cell activation, the formation of TCR nanoclusters would induce aggregation of membrane cholesterol, which could provoke the fluorescence signal of the COOH-TPE-PEG-Chol due to the AIE characteristic. Moreover, the enhancement of the fluorescence intensity was positively related to the activation state of T cells. Our study demonstrated the concept of cholesterol-derived AIE fluorescent probes for deciphering the spatiotemporal arrangements of TCR on the membrane during T cell activation, and consequently provided a novel and complementary strategy for detecting T cell activation in real-time.