Photoimmunotechnology as a powerful biological tool for molecular-based elimination of target cells and microbes,...

Microbial pathogens, including bacteria, fungi and viruses, can develop resistance to clinically used drugs; therefore, finding new therapeutic agents is an ongoing challenge. Recently, we reported the photoimmuno-antimicrobial strategy (PIAS), a type of photoimmunotechnology, that enables molecularly targeted elimination of a wide range of microbes, including the viral pathogen severe acute respiratory syndrome coronavirus 2 and the multidrug-resistant bacterial pathogen methicillin-resistant Staphylococcus aureus (MRSA). PIAS works in the same way as photoimmunotherapy (PIT), which has been used to treat recurrent head and neck cancer in Japan since 2020. Both PIAS and PIT use a monoclonal antibody conjugated to a phthalocyanine derivative dye that undergoes a shape change when photoactivated. This shape change induces a structural change in the antibody-dye conjugate, resulting in physical stress within the binding sites of the conjugate and disrupting them. Therefore, targeting accuracy and flexibility can be determined based on the specificity of the antibody used. In this protocol, we describe how to design a treatment strategy, label monoclonal antibodies with the dye and characterize the products. We provide detailed examples of how to set up and perform PIAS and PIT applications in vitro and in vivo. These examples are PIAS against microbes using MRSA as a representative subject, PIAS against viruses using severe acute respiratory syndrome coronavirus 2 in VeroE6/TMPRSS2 cells, PIAS against MRSA-infected animals, and in vitro and in vivo PIT against cancer cells. The in vitro and in vivo protocols can be completed in ~3 h and 2 weeks, respectively.