Lillian Lee

ATRP-mediated continuous assembly of polymers for the preparation of nanoscale films

The continuous assembly of polymers (CAP) via atom transfer radical polymerisation (ATRP) is reported as an efficient approach for the preparation of dense, cross-linked, nanoscale engineered films as surface coatings, hollow capsules and replica particles. These films can be reinitiated to allow the preparation of thicker films without loss of film growth efficiency while maintaining similar film density.

Manipulating the salt and thermal stability of DNA multilayer films via oligonucleotide length.

DNA films are promising materials for diverse applications, including sensing, diagnostics, and drug/gene delivery. However, the ability to tune the stability of DNA films remains a crucial aspect for such applications. Herein, we examine the role of oligonucleotide length on the formation, and salt and thermal stability, of DNA multilayer films using oligonucleotides of homopolymeric diblocks (polyAG and polyTC), with each block (A, G, T, or C) ranging from 5 to 30 bases (10-, 20-, 30-, 40-, and 60-mer). Using a combination of quartz crystal microgravimetry, dual polarization interferometry, and flow cytometry, we demonstrate that at least 10 bases per hybridizing block in the DNA diblocks (that is, 20-mer) are required for successful hybridization and, hence, DNA multilayer film formation. Films assembled using longer oligonucleotide blocks were more stable in low salt conditions, with the DNA multilayer films assembled from the 60-mer oligonucleotides remaining intact in solutions of about 25 mM NaCl. A systematic increase in film melting temperature ( T m) was observed for the DNA multilayer films (assembled on colloids) with increasing oligonucleotide length, ranging from 38.5 degrees C for the 20-mer films to 53 degrees C for the 60-mer films. Further, an alternating trend in T m of the DNA multilayer films was observed with layer number (AG or TC); DNA multilayer films terminated with an AG layer exhibited a higher T m (44-49 degrees C) than films with an outermost TC layer (ca. 38 degrees C), suggesting a rearrangement of the film structure upon hybridization of the outermost layer. This work shows that the stability of DNA multilayer films can be tuned by varying the length of the oligonucleotide building blocks, thus providing a versatile means to tailor the salt and thermal stability of DNA films, which is necessary for the application of such films.

A Novel Acoustomicrofluidic Nebulization Technique Yielding New Crystallization Morphologies

A novel acoustic microfluidic nebulization platform is demonstrated, which, due to its unique ability to access intermediate evaporation rate regimes-significantly faster than that in slow solvent evaporation but considerably below that achieved in spray drying, is capable of producing novel crystal morphologies that have yet to be reported in both model inorganic and organic systems. In addition, the potential for simultaneously encapsulating single crystals within a biodegradable polymeric coating in a single simultaneous step together with the crystallization process as the solvent evaporates during nebulization is briefly shown. The platform not only has the potential to be highly scalable by employing a large number of these low-cost miniature devices in parallel to achieve industrially relevant particle production rates, but could also be advantageous over conventional spray drying in terms of energy utilization, given the tremendous efficiency associated with the high-frequency ultrasonic microdevice as well as its ambient temperature operation.

Enzyme-Based Listericidal Nanocomposites

Cell lytic enzymes represent an alternative to chemical decontamination or use of antibiotics to kill pathogenic bacteria, such as listeria. A number of phage cell lytic enzymes against listeria have been isolated and possess listericidal activity; however, there has been no attempt to incorporate these enzymes onto surfaces. We report three facile routes for the surface incorporation of the listeria bacteriophage endolysin Ply500: covalent attachment onto FDA approved silica nanoparticles (SNPs), incorporation of SNP-Ply500 conjugates into a thin poly(hydroxyethyl methacrylate) film; and affinity binding to edible crosslinked starch nanoparticles via construction of a maltose binding protein fusion. These Ply500 formulations were effective in killing L. innocua (a reduced pathogenic surrogate) at challenges up to 105 CFU/ml both in non-growth sustaining PBS as well as under growth conditions on lettuce. This strategy represents a new route toward achieving highly selective and efficient pathogen decontamination and prevention in public infrastructure.

Effect of oligonucleotide length on the assembly of DNA materials: Molecular dynamics simulations of layer-by-layer DNA films

DNA strand length has been found to be an important factor in many DNA-based nanoscale systems. Here, we apply molecular dynamics simulations in a synergistic effort with layer-by-layer experimental data to understand the effect of DNA strand length on the assembly of DNA films. The results indicate that short (less than 10 bases) and long (more than 30 bases) single-stranded DNAs do not exhibit optimal film growth, and this can be associated with the limited accessibility of the bases on the surface due to formation of self-protected interactions that prevent efficient hybridization. Interestingly, the presence of a duplex attached to a single strand significantly alters the persistence length of the polyT strands. Our study suggests that restrained polyT, compared to labile suspensions of free polyT, are more capable of hybridization and hence DNA-based assembly.

Influence of salt concentration on the assembly of DNA multilayer films.

DNA multilayer films are promising candidates for a plethora of applications, including sensing, diagnostics, and drug/gene delivery. Fabricated solely from DNA, the use of salt in forming DNA multilayers is crucial in promoting and maintaining hybridization of complementary base pairs by minimizing the repulsive forces between the oligonucleotides and preventing disassembly of the layers once formed. Herein, we examine the role of salt on the assembly of DNA films assembled from oligonucleotides composed of two homopolymeric diblocks (polyA(n)G(n) and polyT(n)C(n)) in salt concentrations ranging from 0.1 to 2 M. Using quartz crystal microgravimetry (QCM) and flow cytometry, we show that films assembled at high salt concentrations (2 M salt) exhibit a different morphology and are denser than those assembled from lower (1 M salt) salt solutions. Formation of the T x A*T triplex in solution and within the DNA film was also studied using circular dichroism (CD) and QCM, respectively. DNA films assembled using oligonucleotides of various lengths (20- to 60-mer) at high salt concentration (2 M salt) showed no significant influence on the film growth. This work shows that salt plays an important role in the assembly and final morphology of DNA multilayer films, hence enabling films with different properties to be tailored.

Probing the dynamic nature of DNA multilayer films using Förster resonance energy transfer.

DNA films are of interest for use in a number of areas, including sensing, diagnostics, and as drug/gene delivery carriers. The specific base pairing of DNA materials can be used to manipulate their architecture and degradability. The programmable nature of these materials leads to complex and unexpected structures that can be formed from solution assembly. Herein, we investigate the structure of DNA multilayer films using Förster resonance energy transfer (FRET). The DNA films are assembled on silica particles by depositing alternating layers of homopolymeric diblocks (polyA(15)G(15) and polyT(15)C(15)) with fluorophore (polyA(15)G(15)-TAMRA) and quencher (polyT(15)C(15)-BHQ2) layers incorporated at predesigned locations throughout the films. Our results show that DNA films are dynamic structures that undergo rearrangement. This occurs when the multilayer films are perturbed during new layer formation through hybridization but can also take place spontaneously when left over time. These films are anticipated to be useful in drug delivery applications and sensing applications.

Programmed degradation of DNA multilayer films.

The design and assembly of DNA multilayer films with programmable degradation properties are reported. The nanostructured DNA films are assembled through the layer-by-layer (LbL) assembly technique and can be programmed to degrade by subsequently introducing DNA strands of specific sequences. The strands preferentially hybridize to the building blocks that stabilize the film structure, causing the film to rearrange and degrade. The rate of degradation is influenced by both the availability and accessibility of the complementary DNA binding sites within the film, as well as the degree of crosslinking within the film. Similar results are obtained for DNA multilayer films assembled on planar and particle supports. This approach offers an avenue to tailor degradability features into DNA-based materials that may find application in the biosciences, in areas such as biosensing and drug delivery.

Hybrid Surface and Bulk Resonant Acoustics for Concurrent Actuation and Sensing on a Single Microfluidic Device

While many microfluidic devices have been developed for sensing and others for actuation, few devices can perform both tasks effectively and simultaneously on the same platform. In piezoelectric sensors and actuators, this is due to the opposing operating requirements for sensing and actuation. Sensing ideally requires narrow resonant peaks characterized by high quality factors, such as those found in quartz crystals. However, these materials usually have poor electromechanical coupling coefficients that are not ideal for actuation. In this work, we show that it is possible to achieve both sensing and actuation simultaneously on a shared device by exploiting the distinct advantages of both bulk waves for effective mass sensing and surface waves for highly efficient microfluidic actuation through a unique hybrid surface and bulk acoustic wave platform. In light of the recent resurgence of interest in portable inhaled insulin devices for personalized diabetes management, we demonstrate the use of this technology for efficient aerosolization of insulin for inhalation without denaturing the protein, while being able to concurrently detect the residual mass of the un-nebulized insulin remaining on the device such that the actual dose delivered to the patient can be determined in real time.