Quantitative evaluation of minimal polynucleotide concentrations has become a critical analysis among a myriad of applications found in molecular diagnostic technology. Development of high-throughput, nonenzymatic assays that are sensitive, quantitative and yet feasible for point-of-care testing are thus beneficial for routine implementation. Here, we develop a nonenzymatic method for quantifying surface concentrations of labeled DNA targets by coupling regulated amounts of polymer growth to complementary biomolecular binding on array-based biochips. Polymer film thickness measurements in the 20-220 nm range vary logarithmically with labeled DNA surface concentrations over two orders of magnitude with a lower limit of quantitation at 60 molecules/μm 2 (∼10 6 target molecules). In an effort to develop this amplification method towards compatibility with fluorescence-based methods of characterization, incorporation of fluorescent nanoparticles into the polymer films is also evaluated. The resulting gains in fluorescent signal enable quantification using detection instrumentation amenable to point-of-care settings.

The reaction and coating kinetics for the glucose oxidase initiated interfacial polymerization are elaborated. The interfacial film grows rapidly and linearly with time, producing time-dependent controllable conformal coating thicknesses of up to a millimeter in less than 4 minutes. Bulk polymerization was only observed when the immersing media was stirred to induce higher mass transport rates. The dramatically different film thicknesses observed between different concentrations of glucose in the hydrogel and iron in the bulk media are demonstrated to be a result of an initial rapid growth phase following which the film grows at the same rate nearly independent of either the glucose or iron concentration. The polymerization rate and hence the thickness growth rate in this initial phase saturate at glucose and iron concentrations above 0.8 M and 0.63 mM, respectively. At iron concentrations above 0.05 mM, the film thickness at the end of 3 hours of reaction monotonically decreased with increasing iron concentration from 5.7 mm to 4.2 mm. The glucose oxidase is trapped by the growing polymerization front and can be used as the sole enzymatic precursor to coat a second polymeric layer. However, the rate of film growth of the second layer is 14-fold lower than the rate of film growth when bulk enzyme is present during the second stage coating process.

Here, we report a novel method of suppressing oxygen inhibition while simultaneously performing photopolymerizations with a visible light photoinitiator. A singlet oxygen sensitizer, Zinc tetrakis(tert-butyl) phthalocyanine (Zn(ttp)), is used to excite oxygen at a rate greatly exceeding the initiation rate to promote the reaction of ground state oxygen preferentially with Zn(ttp). The inhibition times of such polymerizations decreased from 280 s for polymerization without Zn(ttp) to 40 s for Zn(ttp)-mediated polymerizations in air. Thermal polymerization studies shows that the suppression mechanism happens only when the formulation is exposed to visible light that is absorbed by Zn(ttp). The excited state of Zn(ttp) resulting from light absorption transfers its energy to oxygen, consequently exciting it to the singlet state. Because of the presence of deactivating mechanisms, the singlet oxygen decays back to its ground state and a quasi-steady state is established between the ground state and singlet state oxygen species. Because of the high rate of light absorption by Zn(ttp) as compared to the photoinitiator, the singlet state oxygen becomes the predominant oxygen species, thereby resulting in significant suppression of radical scavenging by ground state oxygen. Experimental results are corroborated with a mathematical model to describe the variation in inhibition times with changing Zn(ttp) concentration. The value of the characteristic lifetime of singlet oxygen calculated from the model is 1 ms, consistent with that which is reported in the literature for several organic monomers. Finally, optimal initiating conditions are designed to suppress oxygen inhibition while achieving high conversions in the most challenging condition of a sample equilibrated with a pure oxygen atmosphere.

A unique design paradigm to form core–shell particles based on interfacial radical polymerization is described. The interfacial initiation system is comprised of an enzymatic reaction between glucose and glucose oxidase (GOx) to generate hydrogen peroxide, which, in the presence of iron (Fe2+), generates hydroxyl radicals that initiate polymerization. Shell formation on prefabricated polymeric cores is achieved by localizing the initiation reaction to the interface of the core and a surrounding aqueous monomer formulation into which it is immersed. The interfacially confined initiation reaction is accomplished by incorporating one or more of the initiating species in the particle core and the remainder of the complementary initiating components in the surrounding media such that interactions and the resulting initiation reaction occur at the interface. This work is focused on engineering the reaction behavior and mass transport processes to promote interfacially confined polymerization, controlling the rate of shell formation, and manipulating the structure of the core–shell particle. Specifically, incorporating GOx in the precursor solution used to fabricate cores ranging from 100 to 200 μm, and the remainder of the complementary initiating components and monomer in the bulk solution prior to interfacial polymerization yielded shells whose average thickness was 20 μm after 4 min of immersion and at a bulk iron concentration of 12.5 mM. When the locations of glucose and GOx are interchanged, the average thickness of the shell was 15 or 100 μm for bulk iron concentrations of 45 and 12.5 mM, respectively. The initial locations of glucose and GOx also determine the degree of interpenetration of the core and the shell. Specifically, for a bulk iron concentration of 45 mM, the thickness of the interpenetrating layer averaged 12 μm when GOx was initially within the core, whereas no interpenetrating layer was observed when glucose was incorporated in the core. The polymeric shell formed by this technique is also demonstrated to be self-supporting following core degradation. This behavior is accomplished by fabricating the particle core hydrogel from monomers possessing degradable groups that can be irreversibly cleaved by light exposure following shell formation. When the coated particle was exposed to light, the shell remained intact while the core degraded as evidenced by a dramatic change in diffusion coefficient of fluorescent beads immobilized within the core.

A mathematical model describing interfacial radical polymerization‐based film formation on hydrogels is elucidated. A glucose oxidase‐mediated multistage initiation reaction is used to accomplish interfacial film formation. A polymer concentration‐dependent diffusion coefficient is used to reflect the changing mass transport conditions as the film develops. Model predictions of the film thickness as a function of the species concentrations agree well with experiments. The model predicts that the degree of initiation reaction delocalization with the enzyme‐mediated initiation system is significantly higher than an enzyme‐independent system, thus affecting the film growth rate and structure. The mass transport properties of the film and its adhesion to the underlying substrate are also investigated.