We propose to create a modular suite of products that will make highly multiplexed imaging of individual cells in intact tissue available to a wide range of laboratories through validated labeling kits and companion imaging software that promotes discovery through interactive image and data visualization. Our goal is to create a suite of integrated labeling, display and analysis products to elucidate patterns of disease mechanisms in tissue, which is not possible using current methods. To ensure this goal, we have established a close partnership with Oregon Health Science University (OHSU) and the commitment of a strong group of collaborators in the cancer research community. We have developed a novel cycIF technology based on antibody conjugated oligonucleotides (Ab- oligos). Briefly, a single stranded oligo (docking strand, DS) is conjugated to the primary antibody which can be hybridized to a complementary single stranded oligo (imaging strand, IS) conjugated to a conventional fluorophore. Each DS/IS pair was designed to contain a restriction enzyme (RE) site for selective removal of fluorescence permitting cycles of re-staining and imaging. Currently, little functionality exists to optimally extract the wealth of information in 60+ biomarker channel images. We introduce a novel tool that can dynamically visualize such images, take advantage of the many channels for better cell segmentation even in complex tissue, and automatically correct and register the multiple cycles. In Phase I, a prototype of the imaging system will be built and tested with a subset of labels. Upon confirming feasibility, Phase II will focus on the creation of specific kits and software products in close harmony between biochemistry, software, and research needs.
We propose to create a modular suite of products that will make highly multiplexed imaging of individual cells in intact tissue available to a wide range of laboratories. Our goal is to create a suite of integrated labeling, display and analysis products to elucidate patterns of disease mechanisms in tissue, which is not possible using current methods, which should help lead to more effectively strategies for therapy.
