Controlled trapping and release of quantum dots in a DNA-switchable hydrogel.

DNA-switchable structures have been under extensive investigation in DNA nanotechnology, where DNA-based supramolecular assemblies are switched between mechanically distinct conformational states through hybridization with DNA “fuel” strands. Reversibility is achieved by displacing previously added fuel strands from the assembly in a branch-migration process. This operation principle has been recently applied to realize a switchable polyacrylamide (PAAm) hydrogel with controllable macroscopic rheological properties. In the present study, we investigate the nanoscopic aspects of this system in detail using fluorescent semiconductor quantum dots (QDs) as probe particles. The diffusion properties of the QDs in the gel are studied using single-molecule fluorescence microscopy and fluorescence correlation spectroscopy. Trapping and DNA-triggered release of the nanoparticles is directly visualized, demonstrating the potential of the DNA-switchable gel as a controlled release system with possible applications in drug delivery. PAAm gels are usually synthesized by mixing acrylamide monomers, crosslinkers, and water in a certain ratio, followed by initiation of polymerization with a catalyst. The resulting polyacrylamide chains are chemically inert and biocompatible. Numerous crosslinking agents can be used to tune the properties of PAAm gels. Using double-stranded DNA as a reversible crosslinker provides a number of advantages: the resulting gel is biocompatible and sequence programmable; the mechanical properties and the pore size of the crosslinked gel can be adjusted by the length of the DNA crosslinker strands, and the melting temperature of the gel can be controlled by the sequence and length of the DNA crosslinkers. For the synthesis of the DNA-switchable gel, two noncomplementary Acrydite-modified oligonucleotides, A and B, are separately copolymerized with acrylamide (4% w/v) and thereby incorporated into the PAAm chains. Mixing of the two solutions yields a highly viscous fluid, which, in our experiments, is complemented with fluorescent colloidal semiconductor quantum dots as tracer particles. Fluorescent QDs are easy to track with single-molecule fluorescence techniques and they are available with a wide variety of functionalizations for biomedical applications. Addition of gelation oligonucleotides complementary to the Acrydite strands transforms the fluid PAAm/nanoparticle mixture into a solid gel (Figure 1), trapping the particles. The DNA crosslinker strands are equipped with an additional, unhybridized “toehold” section that acts as a “recognition tag” for DNA release strands. When release strands fully complementary to the crosslinker strands are added to the gel, they attach to the toeholds and remove the crosslinker strands via branch migration. The gel dissolves into a solution, liberating the trapped particles. For a 4% acrylACHTUNGTRENNUNGamide “one-dimensional” (1D) gel, that is, for unlinked linear polymer strands, a typical pore size much larger than the particle diameter is expected, which is considerably reduced by the crosslinking process. The crosslinking density used in our experiments corresponds to a value of %C= 0.16 in a conventional PAAm gel crosslinked with N,N’methylene bisACHTUNGTRENNUNG(acrylamide). Compared to typical polyacrylamide gels used for gel electrophoresis, the DNA–PAAm gel is sparsely crosslinked. For a bis-crosslinked PAAm gel with the same parameters, a pore size of rp>100 nm would be expected. Nevertheless, we experimentally observe that the DNA-switchable gel is capable of controllably trapping and releasing nanoparticles with a diameter on the order of 10 nm. To characterize the trapping and release process in detail, the diffusion behavior of our tracer particles was studied in the sol and gel states of the switchable gel. As tracer particles, we used CdSe/ZnS QDs (lem=565 nm) with a “bare” radius of roughly 6 nm. The QDs were made water soluble by a polymer coating that slightly increased their radius. As described previously, the radius of the coated QDs was determined using fluorescence correlation spectroscopy (FCS). FCS measurements on free QDs in water yielded a diffusion constant of DH2O=29 0.5 mm s 1 (Figure 2b). Using the Stokes–Einstein (SE) equation this value can be translated into a hydrodynamic radius of the particles of 7.4 0.6 nm. We then determined the diffusion properties of the QDs for the two states of the gel using both single-molecule fluorescence microscopy and FCS. In the unlinked hydrogel matrix the diffusion constant dropped to DFCS,unlinked=6.6 0.3 mms 1 (Figure 2b). Again using the SE equation, one can estimate the (local) viscosity of the unlinked gel matrix to be h=4.8 0.5 mPas. Tracking individual QDs in the unlinked gel matrix by fluorescence microscopy (Figure 2c,d) yields a diffusion constant of DTracking,unlinked=6.3 0.4 mms , which agrees well with the [*] Dr. T. Liedl, Dr. F. C. Simmel Department of Physics and Center for Nanoscience Ludwig-Maximilians-Universit&t M'nchen Geschwister-Scholl-Platz 1, 80539 M'nchen (Germany) Fax: (+49)89-2180-3182 E-mail: simmel@lmu.de