Direct visualization of immune status for tumor-infiltrating lymphocytes by rolling circle amplification

The immune status of tumor-infiltrating lymphocytes (TILs) is essential for the effectiveness of cancer immunotherapies. However, due to the diversity of immune status in TILs, cellular heterogeneity, and the applicability to the clinic, it is still lacking effective strategies to meet clinical needs. We developed a novel immuno-recognition-induced method based on rolling circle amplification (RCA), namely immunoRCA, to in situ visualize the immune status of TILs in actual clinical samples. This developed immunoRCA method, in which, feature mRNAs were used as the biomarkers for the immune status of TILs, has a low fluorescence background, high sensitivity, and specificity. The immunoRCA was able to efficiently evaluate the immune status of CD8+ T cells regulated by activating or inhibiting factors, track the T cell type and immune status during in vitro expansion, and in situ visualize the number, location, and immune status of TILs in clinical specimens.