(31f) DNA-Caged Polymer Nanocomposites for Erasable Fluorescence Imaging

Purpose: DNA follows well-known base pairing rules that allow for the creation of novel structures, including DNA origami. DNA origami can be designed into a wide variety of two- and three-dimensional shapes for a variety of applications, such as nanoscale engineering and drug delivery. Here, we report DNA cages, DNA nanostructures designed to form a network on surfaces. This DNA nanostructure was originally designed by Kurokawa et al.¹ for use as an artificial cytoskeleton inside liposomes. As a cytoskeleton, DNA cages increased the mechanical integrity of the lipids. Our goal is to integrate DNA cages with known micelle structures formed from block copolymers for applications in imaging, including erasable labeling of biological samples. Current procedures for multiplexed imaging via erasable labels require harsh treatment of tissue that can damage the samples. Multiplexed images allow for the labeling of multiple targets simultaneously with high image resolution, which means that complicated pathways in samples such as tumors can be studied using less time and materials. If the samples are damaged before all desired parts are labeled, additional samples may be required, which is not always possible when the samples come from a patient. Further, sequential sample labeling precludes evaluating the same markers in the same cells. The proposed labeling system is based on gentle DNA dehybridization interactions, which would greatly improve current technology.

Methods: The DNA origami previously described¹ was introduced to polymer micelles and solid polymer nanoparticles, forming a cage on their surfaces. Previously¹, it was suggested that DNA interacts with the surface of the micelles based on the charged group on the surface of the micelle and the negatively-charged DNA strands. Here, we show that attraction may also be based on the affinity of DNA for the polymer used. Proof of cage formation was measured based on the fluorescence of the cage itself, until a saturation point on the surface of the micelles was reached. DNA cages were tagged with single stranded DNA (ssDNA) targeting strands that reversibly attach to ssDNA strands labeled with fluorophores via strand displacement reactions. Strand displacement reactions are based on the highest number of complimentary base pairs. Thus, when an ssDNA with greater complement was added, the labeling strand was removed, erasing the signal.

Results and Implications: These data suggest that DNA cages have great promise for multiplexed cell labeling. Proper diagnosis of patients, especially when studying tumors, is highly dependent on cell labeling. DNA cages do not require harsh chemicals or procedures for removal, which may improve tissue labeling. The erasable nature of DNA cages allows for the labeling of the same target multiple times or different targets sequentially, potentially without damaging the sample. Finally, when combined with advanced image analysis techniques, it may be possible to create a three-dimensional image of the sample for a more complete analysis. Thus, DNA cages have the potential to significantly change cell labeling, especially that of patient samples.

Reference

1 Kurokawa, C. et al. DNA cytoskeleton for stabilizing artificial cells. Proc Natl Acad Sci U S A 114, 7228-7233, doi:10.1073/pnas.1702208114 (2017).