Search

Abstract Intravital microscopy (IVM) allows spatial and temporal imaging of different cell types in intact live tissue microenvironments. IVM has played a critical role in understanding cancer biology, invasion, metastases, and drug development. One considerable impediment to the field is the inability to interrogate the tumor microenvironment and its communication cascades during disease progression and therapeutic interventions. Here, a new implantable perfusion window chamber (PWC) is described that allows high‐fidelity in vivo microscopy, local administration of stains and drugs, and longitudinal sampling of tumor interstitial fluid. This study shows that the new PWC design allows cyclic multiplexed imaging in vivo, imaging of drug action, and sampling of tumor‐shed materials. The PWC will be broadly useful as a novel perturbable in vivo system for deciphering biology in complex microenvironments.

Abstract The ability to observe cells in live organisms is essential for understanding their function in complex in vivo milieus. A major challenge today has been the limited ability to perform higher multiplexing beyond four to six colors to define cell subtypes in vivo. Here, a click chemistry‐based strategy is presented for higher multiplexed in vivo imaging in mouse models. The method uses a scission‐accelerated fluorophore exchange (SAFE), which exploits a highly efficient bioorthogonal mechanism to completely remove fluorescent signal from antibody‐labeled cells in vivo. It is shown that the SAFE‐intravital microscopy imaging method allows 1) in vivo staining of specific cell types in dorsal and cranial window chambers of mice, 2) complete un‐staining in minutes, 3) in vivo click chemistries at lower (µm) and thus non‐toxic concentrations, and 4) the ability to perform in vivo cyclic imaging. The potential utility of the method is demonstrated by 12 color imaging of immune cells in live mice.