A Fluorescent label for 3D optical imaging of administered substances in tissues

Abdullah Syed 1 ;  Jessica Ngai 1, 3 ;  Presley MacMillan 4 ;  Stefan Wilhelm 6 ;  Shrey Sindhwani 1 ;  Benjamin Kingston 1 ;  Warren C.W. Chan 1, 2, 3, 5

1. Institute of Biomaterials and Biomedical Engineering, University of Toronto, 164 College St, Toronto, ON, M5S 3G9, Canada; 2. Donnelly Center for Cellular and Biomolecular Research, University of Toronto, 160 College St, Toronto, ON, M5S 3E1, Canada; 3. Chemical Engineering and Applied Chemistry, University of Toronto, 200 College St, Toronto, ON, M5S 3E5; 4. Department of Chemistry, University of Toronto, 80 St. George, Toronto, ON, M5S 3H6; 5.  Materials Science and Engineering, University of Toronto, 14 College St, Toronto, ON, M5S 3G9, Canada;  6.  Stephenson School of Biomedical Engineering, University of Oklahoma, 101 David L. Boren Boulevard, Norman, Oklahoma 73072, United States

3D microscopy in whole organs and animals can provide unprecedented insight into biological structure and function. This is achieved by chemically treating tissues with solvents or detergents to remove lipids and make the tissue transparent. However, these approaches are limited in their ability to visualize injected substances because the chemical treatment process required to prepare tissues for 3D microscopy removes any uncrosslinked molecules. Here, we demonstrate the imaging of small molecules, lipids, polymers and carbohydrates by labelling them with a fluorescent peptide tag that can crosslink into tissue. We developed a simple and reproducible reaction scheme using maleimide-cysteine conjugation to allow facile labelling of most biochemicals with this tag. We found that this tag could be readily visualized throughout the body even during its rapid transit through renal tubules. We also managed to retain organic nanoparticles in tumour tissue in 3D and were able to distinguish between intact and degraded liposomes by using one tag for the lipids and another tag for the encapsulated volume and examining the 3D distribution of each. This simple labelling technique opens the way to investigating a wide range of biological processes that depend on the transport of administered substances.