Abstract Cancer immunotherapy is revolutionizing the landscape of cancer treatment, with checkpoint inhibitors, activated T cell therapies, and a range of other combinatorial approaches providing lasting benefits for patients. However, the response rate to these advanced therapeutic approaches is only 15-30%, demonstrating the urgent need of better understanding of and eventually addressing the low-response problem, as well as the need of technologies for patient stratification, which often involves the use of biomarkers to fingerprint individuals’ tumors. Here, we propose to develop a versatile quantum dot (Qdot)- based single-cell molecular profiling technology for in situ multiplexed quantitative molecular characterization of biological specimens with optical imaging resolution. Despite the availability of Qdots commercially since early 2000s, they have not received broad adoptions and enabled biological discoveries, although they offer much better optical properties than organic fluorophores. Key problems include the modest improvement in multiplexing, and the sophisticated, lengthy, cost-prohibitive, and low-reproducibility probe preparation approaches developed to date. Based on a breakthrough made by Prof. Gao’s lab (Nat. Commun.), the technology proposed here is capable of solving both problems (patent recently awarded, ready for commercialization). It simultaneously achieves the multiplexing capability of mass spectrometry (10s-100s) and the sensitivity, resolution, in situ imaging, low cost, great accessibility (microscopes are standard compared to mass spectrometers) of immunofluorescence.