In this study, we describe an efficient approach to identify and isolate tumour-reactive TCRs based on a combination of T cell activation markers and effector molecules (IFNG, IL2, TNF, IL2RA, CD69, TNFRSF9, GZMB, GZMA, GZMK, and PRF1) through single-cell sequencing. The gene discussed is PRF1; the disease is neoplasm.