Chapter 6: Fabrication of nanostructures using natural synthesis: optical materials using silk – Optical Biomimetics

6

Fabrication of nanostructures using natural synthesis: optical materials using silk

H. Tao, D.L. Kaplan and F.G. Omenetto,     Tufts University, USA

Abstract:

Biopolymers are promising building blocks for a new generation of green devices. Silk proteins represent a unique family of biopolymers due to their novel structural and biological properties, which serve as a broad inspiration to develop biological foundries for technological applications that leverage nature’s materials as their main constituents. This chapter focuses on the opportunities offered by the material as a promising biopolymer platform for high technology applications in optics and photonics, electronics and optoelectronics.

Key words

silk

biopolymer

photonics

electronics

sustainable materials

6.1 Introduction

The possibility of adopting sustainable materials as alternatives to contemporary technological materials whose fabrication is based on non-renewable resource consumption such as oil, coal or natural gas is a global challenge. Naturally occurring materials provide a compelling template to reinterpret and, in some cases, simplify modern manufacturing while at the same time rendering it sustainable.

A particular and long-standing challenge is to identify, among the options available, new materials that, while being environmentally sustainable, also maintain adequate physical and material properties to meet the performance requirements and the fabrication tolerances needed successfully to interface with current technology. Further, such materials have to be widely available and be cost-competitive within the global commodity supply chain. A conjunction of practical and technological elements is needed to make naturally occurring materials a credible alternative to current plastic, semiconductors and inorganic substrates, and provide alternatives to reroute present-day materials towards more environmentally sustainable options.

Biopolymers are promising building blocks for a new generation of green devices. This class of polymers, produced and modified by living organisms, are the mainstay of naturally occurring, self-assembling, structurally hierarchical micro- and nanoscale systems, from the polysaccharide chitin in butterfly wings and beetle exoskeletons, to the collagen in lens arrays or dermal iridescences, to the keratin in peacock feathers, among many others. The direct utilization and re-engineering of these biological materials into technological material platforms is already underway in many laboratories and offers a path forward for a green revolution (Vukusic, Sambles et al. 1999; Sarkar and Mallick 2000; Vukusic, Sambles et al. 2000; Aizenberg, Tkachenko et al. 2001; Sundar, Yablon et al. 2003; Aizenberg 2004; Payne, Yi et al. 2005; Hooper, Vukusic et al. 2006; Steckl, Hagen et al. 2006; Steckl 2007; Yu, Li et al. 2007; Arockiados, Xavier et al. 2008; Cremona, Legnani et al. 2008; Oksman, Mathew et al. 2009; Payne, Meyer et al. 2009; Vukusic, Kelly et al. 2009; Singh, Sariciftci et al. 2010; Irimia-Vladu, Sariciftci et al. 2011).

We will focus on the opportunities offered by silk proteins as a promising biopolymer platform for high technology applications (Vollrath and Knight 2001; Shao, Vollrath et al. 2003; Shao, Yang et al. 2005; Shao, Chen et al. 2006; Omenetto and Kaplan 2008; Vollrath, Liu et al. 2008; Young, Brookes et al. 2008). In this chapter, we will address the use of silk proteins in optics and photonics, electronics and optoelectronics. These options represent additional developments for this technology platform that compound the broad utility and impact of this material for medical needs that have been recently described in the literature (Cannas, Santin et al. 1999; Motta, Migliaresi et al. 2002; Kaplan, Altman et al. 2003; Kaplan, Meinel et al. 2005; Scheibel 2005; Kaplan, Wang et al. 2006; Kaplan and Vepari 2007; Kaplan, Kluge et al. 2008; Kundu, Dash et al. 2008; Chen, Zhou et al. 2009). The favorable properties of the material serve as a broad inspiration to develop biological foundries for technological applications that leverage nature’s materials as their main constituents.

Silk fibers have historically been highly favored in the textile industry for a few thousand years owing to their extraordinary mechanical strength plus their exceptionally smooth and gleaming appearance. In addition to the popular use in clothing, silk has recently found its applications as a promising biomaterial because of several desirable properties. In fact, silk has been used as a suture material for centuries. From a materials science perspective, silks spun by spiders and silkworms represent the strongest and toughest natural fibers known and offer unlimited opportunities for functionalization, processing, and biological integration. As shown in Table 6.1, the toughness of silk protein fibers, for example spider silks and silkworm silks, is greater than most synthetic materials, including widely used high-performance Kevlar fibers (Vepari and Kaplan 2007). In terms of strength, silk fibers (0.6-1.1 GPa) are in orders of magnitudes higher than that of commonly used polymeric biomaterials such as poly(L-lactic acid) (PLA) (28-50 MPa) and collagen (0.9 –ha 7.4 MPa) (Omenetto and Kaplan 2010). Silks are produced by several kinds of insects (over tens of thousand species) that include, for example, spiders from the class Arachnida, mites, butterflies and moths from the order Lepidoptera. Silk fibers generated from silkworms, known as the Bombyx mori, are of particular interest because of the feasibility of large-scale cultivation (thus cost-effective) and slightly lower but comparable properties to spider silks.

Table 6.1

Mechanical properties of spider and silkworm silks and other materials

The B. mori silk fibroin fibers are typically about 10–25 μm in diameter, consisting of two proteins in a 1 : 1 ratio, i.e. a light chain (~ 26 kDa) and a heavy chain (~ 390 kDa), which are linked by a single disulfide bond and coated with adhesive and hydrophilic sericin proteins (20–310 kDa) (Kaplan, Altman et al. 2003). Silk fibroin is a block copolymer rich in hydro-phobic beta sheet forming protein blocks that self-assemble to form strong and resilient materials with high mechanical strength and toughness. In addition to the notable mechanical properties, silk fibroin is also a degradable and resorbable material. B. mori silk fibroin is reported to be biocompatible and to induce minimal inflammatory responses when implanted (Kaplan, Altman et al. 2003). However, the presence of sericin may cause immunological and allergic reactions in humans, and the removal of sericin from the raw silk cocoons is essential for further medical applications, which can be done by boiling silk cocoons in an alkaline solution, the so-called de-gumming process. Once the sericin proteins are removed, the fibroin fibers can be dissolved into an aqueous solution of pure silk fibroin protein, which can be further processed into different materials ranging from gels and sponges to blocks and films (Rockwood, Preda et al. 2011). Silk films are of particular interest for optics and electronics applications because of their optical transparency (~ 92% across the visible range with the film thickness between 40 μm to 100 μm) and surface smoothness (with a root-mean-square surface roughness of just a few nanometers) (Omenetto and Kaplan 2008). The refractive index of silk (n = 1.54 @ 633 nm) is similar to that of glass (n = 1.52) and higher than that of water (n = 1.33), which implies the possibilities for bio-optics applications, for instance, guiding light within a water-based biological environment with silk waveguides (Omenetto, Parker et al. 2009).

Silk proteins represent a unique family of biopolymers due to their novel structural and biological properties. Recent progress has been made into the relationship between structure and processing. This includes the role of self-assembly (Kaplan, Valluzzi et al. 2002; Braun and Viney 2003; Kundu, Khire et al. 2010), the role of water in assembly and processing (Jin, Park et al. 2005; Lee, Min et al. 2006; Vollrath and Porter 2009; Omenetto, Tsioris et al. 2011), and various options to modify the native proteins (Mori and Tsukada 2000; Goldsmith, Shimada et al. 2005). This has led to the transformation of this ancient and commodity material, in particular silkworm silk, into a variety of new material formats including, hydrogels, ultrathin films, thick films, conformal coatings, three-dimensional (3D) porous or solid matrices, fibers with diameters spanning the nano- to the macroscale, and many related material formats (Fig. 6.1). In addition, silk is processed in an all water-based, room-temperature, neutral pH environment, is mechanically stable, edible, biocompatible, and implantable in the human body.

6.1 Various silk-based materials formats and devices cover a multitude of material scales with sizes relevant to applications covering a wide range of the electromagnetic spectrum from radio frequency (RF) to optical regimes for novel biomedical, optoelectronic, and photonic applications.

What particularly distinguishes silk from other biopolymers for high technology applications are the robust mechanical properties, the facile control of materials properties through the control of water content during processing, the programmable/controllable (from instantaneous to years) degradation lifetime, and the unique optical and electronic properties of the material. Further the ambient environment during silk processing allows for the incorporation of labile biological components without loss of function and with retention of bioactivity over extended time frames (Kaplan, Lu et al. 2009; Kaplan, Lu et al. 2010).

This feature, in particular, provides the opportunity to embed biochemical functions within the biopolymer-based material system, and enables the direct incorporation of biological activity in a bulk material substrate. The latter offers a paradigm shift from traditional optoelectronic and electromagnetic devices that are commonly manufactured on biologically inert substrates, opening opportunities for sustainable and ‘environmentally interactive’ devices.

Silk-based materials appear to be suited to ideally interface with both technological and natural worlds. One of the unique traits of these materials is the ability to store functional compounds, including both organic and inorganic. Since the novel block copolymer structure of silk protein chains provide the template to maintain and preserve biochemical function of such labile biological components (e.g. enzymes, antibodies) within the material structure, new opportunities become available to affect device behavior and responses. The coexistence of technological devices and biological components to generate ‘living photonic’ components (or more generally, ‘living materials’) through the combination of nanofabrication, diffractive optics, and biological doping provides innovative venues for functional devices that are not easily (if at all) attainable with more traditional approaches. This confluence of material properties and functions is the driver for using silk for applications in photonics, electronics, and optoelectronics.

6.2 Silk optics and photonics

Silk has recently emerged as a highly promising material platform for optics and photonics applications because of its excellence on mechanical robustness, optical transparency, and surface flatness. Realization of silk devices starts from production of the silk fibroin solution that is regenerated and purified from natural silk cocoons. The clear water-like silk fibroin solution can be activated, biochemically and/or physically, by simply mixing with various organic (such as cells, proteins, and enzymes) and/or inorganic (such as quantum dots, laser dyes, and metallic nanoparticles) dopants into the solution. The either undoped or doped solution can be deposited on appropriate substrates (flat or patterned), which crystallizes through protein self-assembly upon exposure to air, without the need to resort to exogenous cross-linking reactions or post processing cross-linking for stabilization. This offers the opportunity for numerous fabrication strategies that yield a class of optical elements or a mechanically robust, biocompatible and bioresorbable substrate for thin film photonic and electronic devices (Fig. 6.2).

6.2 Different fabrication strategies for silk devices using silk solution derived from natural silk sources (such as cocoons, webs, and fibers) as the starting material.

Silk fibroin can be easily formed into mechanically robust films of thermodynamically stable beta sheets, with the ability to control thicknesses from just below ten nanometers to hundreds of micrometers or more (Fig. 6.3).

6.3 Control of the thickness of spin-coated silk film through the adjustment of the silk concentration (6 wt% is shown) and spin rate allowing for ultimate film thicknesses down to less than 10 nanometers.

The resulting hardened silk has mechanical properties, surface quality (surface roughness root mean square (rms) < 5 nm) and transparency (over 90% transmission across the visible range) that are ideally suited for its use as an optical substrate (Kaplan, Lawrence et al. 2009). Owing to the processing and self-assembly attributes, silks are suitable for soft-lithography. By casting the aqueous solution on patterned surfaces the topographies can be replicated down to feature sizes less than 10 nanometers in the hardened, free-standing silk film. Using this approach in combination with the optical transparency of the films, a variety of photonic structures can be fabricated. These materials and devices include one-dimensional (1D) and two-dimensional (2D) diffractive structures, holograms, prisms, photonic lattices, and microlens arrays (Fig. 6.4).

6.4 Examples of silk-based optical and photonic elements and devices: (a) an image through a 1-cm-diameter lens; (b) an image through a 6 x 6 microlens array; (c) image of a silk microprism array (scalebar = 25 micrometers); (d) 2 cm long inkjet printed silk waveguide; (e) a scanning electron microscope (SEM) image of a periodic nanohole array with lattice constant equal to 400 nm (scalebar = 500 nm); (f) periodic nanoarrays imprinted in silk (such as in (e)) under dark-field illumination showing different colors associated to the different lattice constants.

While soft-lithography is effective, nanoimprinting of silk films is possible by adjusting their glass transition temperature, which depends on the water content and absorbed moisture of the films themselves (Agarwal, Hoagland et al. 1997). For films prepared at ambient humidity (e.g. ~ 35%) the glass transition temperature is ~ 100°C. This allows nanoimprinting by using a hot embossing process in which a silk film is pressed (~ 30-50 psi) onto a heated (100°C) master with micro- or nanoscale patterns for a few seconds and after a brief (~ 1 min) cooling step can then be mechanically removed from the master (Mondia, Amsden et al. 2010). In contrast, progressive water saturation of the films decreases the glass transition temperature to room temperature.

This allows the additional opportunity to carry out the nanoimprinting process at room temperature by adding a small amount of water on the film surface before the master is pressed onto the silk film (Omenetto, Amsden et al. 2010) (Fig. 6.5). Once imprinted, the films can be annealed by exposure to water vapor or methanol to ensure their insolubility in water (Cao, Lv et al. 2005; Jin, Park et al. 2005; Wen, Xu et al. 2007). Upon exposure to methanol, the secondary structure of the silk fibroin protein changes from (primarily) random coil to (primarily) β-sheet, resulting in an increase in the glass transition temperature (Cebe, Hu et al. 2006). After annealing, the imprinted films are stable and can last for years under ambient conditions. The ability to adopt nanoimprinting techniques increases the quality and throughput of nanostructured silk materials in comparison to casting methods, as well as eliminating mechanical stresses when separating hardened silk from masters (Omenetto, Perry et al. 2008). In addition to the speed and fidelity of production of such films, as well as their mechanical and optical properties, these silk optical devices possess optical features and performance that exceed those of other biopolymers and biocompatible polymers such as chitosan (Park, Cheng et al. 2007) and polylactide (PLA) (Mills, Navarro et al. 2006).

6.5 Scheme illustrating rapid nanoimprinting in silk and resulting images corresponding to the imprinting of a periodic nanohole array.

6.2.1 Silk photonic lattices

The effectiveness of both soft-lithography and nanoimprinting processes replication geometries and topologies down to tens of nanometers allows for more sophisticated nanoscale photonic components to be generated from silk. Of particular interest is the ability to define ordered arrays of deterministic nanoscale features which confer predetermined scattering properties to these protein films, ultimately resulting in color localization when illuminated by white light. Light scattering in periodic systems has been investigated for decades in optics and photonics (Elachi 1976). Their classical description relies on Bragg scattering phenomena, which give rise to constructive interference at specific wavelengths along well-defined propagation directions, depending on illumination conditions, structural periodicity, and the refractive index of the surrounding medium (Baryshev, Khanikaev et al. 2009).

Structural color based on periodic nanopatterned two-dimensional (2D) lattices in silk protein films were demonstrated by imprinting periodic lattices of nanoscale holes (200 nm diameter, 40 nm in depth) which display different colors as a function of varying lattice spacing under white light illumination (Amsden, Perry et al. 2009; Omenetto, Amsden et al. 2010) as shown in Plate XVI (see color plate section between pages 96 and 97). The variation of the index of refraction contrast between the imprinted nanopatterned lattices and the surrounding environment causes a shift in the spectral response as a function of the perturbation.

Plate VII (top) Image of the patterned, undoped silk film illuminated at grazing angle under white light illumination. Different colours appear because of the pitch of the individual patterns. The scheme of the experimental layout is illustrated in the figure as well, whereas the bottom image shows the nanopatterned doped silk film. The enhancement in emission corresponds to specific lattice constants that are matched to the fluorescence spectrum from different pitch nanopatterns. Shown underneath the fluorescent image are images collected at λ = 550 nm and λ = 630 nm, illustrating the specific nanopatterned squares that give rise to enhancement at those wavelengths.

Plate XVI Various periodic lattices imprinted in silk and the associated diffracted spectral signatures as observed under dark-field illumination microscopy.

Plate XVII Scales of P. sesostris viewed from above (a) and below (b), finally revealing their iridescence.

This phenomenon can be exploited by applying liquids on the lattices to monitor their optical properties through the spectral responses (Altug, Adato et al. 2009). Such approaches have been used as the optical transduction mechanisms upon which to build colorimetric sensors and have been demonstrated in a range of materials (Nuzzo, Yao et al. 2010). Specifically for silk, this response was observed by using a nanopatterned lattice and exposing it to a variety of glucose concentrations (Amsden, Perry et al. 2009). Measurement of the spectral responses offered a demonstration of the adequacy of silk fibroin photonic lattices to act as an optical sensing substrate. The unique attribute of biological stabilization is a technologically enabling feature of the silk matrix, which can be coupled to the nanopatterned and optical features of the silk to function as transducers, monitors or enhancers of the entrained biological functions. This level of functionality is not commonly available in current technological materials without complex chemistries or modifications.

An example of this type of device was demonstrated by transforming a simple optical component into an interactive optical component. For these experiments, human hemoglobin was mixed with silk solution and then either cast (Omenetto, Lawrence et al. 2008) or imprinted (Omenetto, Amsden et al. 2010) to form a blood-based diffraction grating (Fig. 6.6). The hemoglobin contained in the free-standing silk film remained active, responding to the oxygenation state of the external environment. This silk-blood-grating was used as the building block of a simple spectrometer, using the diffractive properties of the doped silk as the transduction mechanism for the oxygenation levels of the entrained hemoglobin through its spectral response or, in other words, the grating senses its material constituent. The hemoglobin remains active in the silk-grating system after the device is stored for several months at ambient conditions (Omenetto, Domachuk et al. 2009). The ability to preserve the function of biological dopants within silk films was also demonstrated with the entrainment of enzymes such as glucose oxidase, lipase and peroxidase, where activity was preserved for over a year even at 37°C (Kaplan, Lu et al. 2009; Kaplan, Lu et al. 2010).

6.6 Hemoglobin-doped silk grating acting as a bio-active oxygen sensor.

This outcome is in contrast to stabilization of these enzymes only for a few days in similar conditions outside of the silk matrix (Rao, Gouda et al. 2003). To achieve increased sensitivity in the optical response of the silk nanostructured devices, different lattices can be engineered by nanoimprinting aperiodic structures (Fig. 6.7). The colorimetric fingerprints of these naturally inspired geometries result from multiple scattering of the various spectral components of white light interacting with the nanostructured silk film surfaces in combination with the broadband scattering characteristic of aperiodic systems (Boriskina, Lee et al. 2010; Omenetto, Lee et al. 2010). These unique scattering characteristics of deterministic aperiodic scattering systems and the sensitivity of the associated colorimetric fingerprints to morphological and refractive index surface variations suggest a very appealing path towards the engineering of novel sensing platforms for label-free optical detection of thin molecular layers, all based on this unique pure protein system. Silk films imprinted with such geometries generate complex signatures that are typical of light localization, underscoring the effectiveness of this biopolymer as a technological substrate for advanced photonics.

6.7 A dark-field image of an aperiodic lattice of nanoholes imprinted in silk to obtain complex colorimetric signatures. The inset shows an SEM image of the nanoholes in silk. The entire lattice covers a 50 μim × 50 μim area on a silk film.

6.2.2 Silk-based optical fibers

The same silk solution that is used as the starting material to manufacture the optical films described above, can also be used as a material for micro-fluidic printing. Given the favorable material properties, the use of silk-based inks can be important for a variety of controlled biomaterial fabrication on the microscale. Among the possible applications of this approach is the opportunity to directly print optical devices. This was recently explored by using direct inkjet writing to fabricate silk optical waveguides (Omenetto, Parker et al. 2009). In the demonstration, silk fibroin inks were concentrated (aqueous solution of 28–30 wt% silk, compared to the 6–8 wt% concentration used for films in soft-lithography and nanoimprinting) to obtain the viscosity needed for high-resolution and high-quality structures. The printed silk waveguides retained rod-like morphology by crystallization in a methanol-rich coagulation reservoir. Both straight and curved silk waveguides were generated and guided light, with measured losses ranging between 0.25 dB/cm and 0.81 dB/cm for the straight and curved waveguides, respectively. In this case as well, the utility of this method can be enhanced by leveraging the ease of functionalization offered by the aqueous silk solution. Doped waveguides were generated by mixing a laser dye (Rhodamine 6G) into the silk solution prior to printing. These doped waveguides exhibited significant fluorescence while guiding λ = 532 nm light and were measured to have less losses (~< 0.1 dB/cm) than their undoped counterparts. The devices realized in this fashion are equally amenable to nanoimprinting, which could prove to be valuable for facile generation of distributed feedback lasers or fiber-based biophotonic components.

6.2.3 Fluorescent silk with localized emission enhancement

The utility of nanoimprinting processes to generate silk films with complex photonic lattices can be further enhanced by similarly simple ways to functionalize the silk solution through the addition of organic and inorganic dopants that are easily mixed into the silk solution before forming the photonic elements. This opens the option to seamlessly combine nanoim-printed lattices on doped silk films, thereby consenting to design the interactions between the bulk optical properties (induced by the dopants) and the imprinted optical structures.

This was demonstrated by imprinting photonic lattices on fluorescent silk substrates (Mondia, Amsden et al. 2010). Directional and wavelength-specific fluorescent enhancement was obtained from silk films doped with fluorophores when nanoimprinted with periodic lattices that were designed to have suitable lattice constants and, consequently, suitable optical properties to match the spectral response of the compounds embedded in the bulk film (see Plate VII). The aqueous environment afforded by the silk solution makes it possible to easily incorporate organic and/or inorganic dyes used in biomedical applications. Such materials have the potential to be used as sensors, spectral filters, or light emitting devices and open new opportunities in research for biologically based or biologically active photonic crystals or optical resonators.

6.2.4 Silk-based microfluidics and optofluidics

The ability to reform the material into thick films enables the use of silk as a substrate for microfluidic devices. This exemplifies another potential offered by this biopolymer. Constituting such devices out of a material that is biocompatible and interfaces favorably with a biological environment presents additional opportunities over traditional substrates used in micro-fluidic devices such as siloxane-based elastomers or glass (Borenstein, Bettinger et al. 2007).

The functionality of silk can be increased by chemical modifications. This is generally achieved by exploiting the chemical bonds available within the fibroin structure, namely the acid side chains (~ 1% of the total amino acids) or the tyrosine residues (> 5% of the total amino acids) (Kaplan, Wang et al. 2008). A typical modification used generally relies on azo-benzene molecules. These molecules have been widely used in a variety of applications ranging from cell culture applications (Kaplan, Wang et al. 2008), chemical dyes, pH indicators in a fluid environment (Carofiglio, Brigo et al. 2008), for optical sensors (Carofiglio, Fregonese et al. 2006), and in combination with Raman spectroscopy (Wang and Callahan 1998). In addition, azo-benzene has been studied extensively in materials science and engineering as an optically addressable substrate for holographic data storage (Psaltis and Burr 1998) exploiting the cis-trans isomerization of the molecule (Barrett, Mamiya et al. 2007).

With this in mind, chemically activated microfluidic devices can be manufactured uniting the utility of facile chemistry to the convenience of a fluidic platform that has garnered much attention for convenience of use in diagnostics. Hybrid devices combining the azo-benzene functionality within modified silk films and PDMS microfluidics were shown to generate pH-reactive platforms (Tsioris, Tilburey et al. 2010).

6.3 Silk electronics and optoelectronics

The use of silk for in vivo electronic device applications builds on an extensive history of biocompatibility based on biomedical and tissue engineering studies. This history starts with eons of use of silk as medical sutures, which are FDA approved, and has progressed in recent years to utility in the formation of many human tissues and in vivo studies with silks in various forms, from fibers and films to gels and sponges (Kaplan, Altman et al. 2003; Kaplan and Vepari 2007).

6.3.1 Silk as an electronic material

While the mechanical robustness, transparency, and surface flatness are compelling features for photonics applications, such material properties are also desirable for metal deposition for microelectronic applications. Silk films are insulating and could provide a sustainable alternative to thin dielectric layers used in microelectronics or as supports in microelectronics.

Silk biomaterials have the convenience of possessing material attributes that make them ideal constituents of technological devices such as planar microelectronic structures. Recently, there have been reports of using silk as a low-cost e-paper device (Hwang, Wang et al. 2011). Silk can be used as a replacement for traditional inorganic oxide layers such as SiO2 or poly(methylmethacrylate) (PMMA) (Hwang, Wang et al. 2011; Muller, Hamedi et al. 2011; Omenetto, Capelli et al. 2011; Wen, Yao et al. 2011). Organic thin film transistors (OTFTs) have been manufactured on a glass/ ITO substrate by stacking 400 nm thick silk layers as the gate dielectric, n- and p-type organic semiconductors (N,N’-ditridecylperylene-3,4,9,10-tetracarboxylic diimide (P13), and α,ω-dihexyl-quaterthiophene (DH4T) respectively), and gold gate and source electrodes (Fig. 6.8). Both P13 and DH4T are well-characterized benchmark materials for organic electronics production and provide a suitable test for the electronic performance of silk.

6.8 Silk-based organic field effect transistor and corresponding electrical measurement when using an n-type semiconducting material.

The reported silk dielectric constant for this device configuration is εs = 6 with charge mobility values of μn= 4 × 10− 2 cm2/Vs for the silk-P13 and μp = 1.3 × 10− 2 cm2/Vs for the silk-DH4T. The comparison of the parameters of silk-based devices and the equivalent PMMA and SiO2-based devices reveals that measured voltage thresholds for the silk-P13 transistor is one order of magnitude lower, namely 1.8 V at 20 V, than the respective standard organic field effect transistor (OFET). Additionally, the on/off switch ratio was found to be ~ 104, a value that matches the highest value reported for P13 and DH4T-based thin film organic transistors.

6.3.2 Silk-based electromagnetic metamaterials

Patterning metals on silk has been recently demonstrated by a shadow masking deposition method (Zhang, Tao et al. 2010). Micro-fabricated stencils are used to deposit metals onto silk films in a dry, chemical-free environment, preventing chemical contamination that might be involved in other photolithography-based metal patterning methods such as lift-off processes and wet etching. With this approach, deposition of microscale resonators was demonstrated on silk films. Additionally, this approach maintains the integrity and biocompatibility of the silk films making the resulting materials suitable for incorporation into biological environments such as the human body.

Alternatively, a simpler fabrication technique where a single step transfers metal micro patterns to free-standing silk films under ambient processing conditions has been demonstrated (Omenetto, Tsioris et al. 2011). In this approach, standard photolithography is used to define the metal structures patterned onto pre-silanized hydrophobic silicon wafers. Once this is performed, the silk solution is poured onto the wafer and dried at ambient conditions. During the drying process the silk binds to the metallic pattern on the surface of the silicon wafer resulting in the transfer of the metal patterns from the wafer onto the silk film. This approach is particularly appealing because it allows the parallel fabrication of microstructures over a large area, free-standing and flexible silk films with high precision and eliminating the need for alignment.

The properties of silk allow for the manufacturing of high-quality structures that combine the features of the biopolymer and the capacities offered by microfabrication techniques. With these approaches, large area metamaterial structures were patterned on free-standing silk substrates and exhibited strong resonance responses at Terahertz frequencies. The structures can be scaled to cover a broad area of the electromagnetic spectrum, in spite of the interest provided by the THz regime where numerous chemical and biological agents show unique ‘fingerprints’, which could potentially be used for identification and bio-sensing (Funk, Barber et al. 2005).

The canonical sub-wavelength metamaterial element is the split ring resonator (SRR) (Pendry, Holden et al. 1999). The electromagnetic responseoriginates from oscillating electrons in highly conducting metals such as gold or copper, allowing for a tailored electromagnetic response. The SRR can be thought of as an LC resonator in a simple representation with a resonance frequency of ω0~(1/LC)1/2, where the inductance results from the current path of the SRR and capacitance is mainly determined by the split gap (O’Hara, Singh et al. 2008; Omenetto, Tao et al. 2010b). Any change in the capacitance or the inductance will result in a change in the resonant response, making metamaterials sensitive to the local environment. Therefore, metamaterials would offer an interesting option for integration with silk films for novel sensing and detection applications by providing a natural and biologically active material as the support matrix for the SRR. This was demonstrated by monitoring the evaporation of residual water in five minute intervals from the hardened silk film when the silk film was placed in a dry box, providing access to a new transduction mechanism to monitor hydration state and conformational changes of the fibroin proteins transitioning from amorphous to the crystalline state (Zhang, Tao et al. 2010) (Fig. 6.9). The possibility of scaling electromagnetic structures to cover different electromagnetic regimes and of silk films to be functionalized with biological components offers extensive options for biocompatible sensors, transducers and biologically integrated monitors.

6.9 Shadow masking patterned micro split ring resonators on silk substrates showing strong resonant responses at terahertz frequencies that can be used to monitor the crystallization level of the silk protein.

An additional trait of silk films is the possibility of tuning the properties, such as solid-to-gel-to-liquid state, with stable or dissolvable features. Such tenability in material features can be attained through the control of hydration and the degree of crystallinity (beta-sheet content) in the final material. By leveraging this property, flexible electrodes that conform to curvilinear surfaces are possible (see Fig. 6.10). Electronics that are capable of intimate, noninvasive integration with the soft, curvilinear surfaces of biological tissues offer important opportunities for monitoring, diagnosing and treating disease or injury, and for establishing brain/machine interconnects.

6.10 A ~ 1 cm × 1 cm electrode array on a dissolvable silk film conformally transferred/wrapped on a rounded surface.

6.3.3 Silk-integrated plasmonics

In addition to the ability of being reshaped with complex nano- and microscale topologies, silk provides a suitable interface for microelectronic and photonic applications. This allows the use of silk for conformal coatings on a variety of materials (hydrophobic, hydrophilic, plastic, metal, among others) with control across various length scales, ranging from monolayers to the mesoscale. Thin layers of silk can be obtained by spin coating, dip coating or spray coating the solution. Solution concentration, additives, and application rates can be used to modulate coatings characteristics.

In early work we showed that sub-nanometer coatings of silk could be conformally deposited by dip coating, using a layer-by-layer approach (Kaplan, Wang et al. 2005; Kaplan, Wang et al. 2007). The concentration of silk in aqueous solution and the presence of salts were used to regulate layer thickness with precision. Similar variables were used to control spin coating of silk solution to generate precisely tunable layers of conformal coating on a variety of surfaces.

This degree of control allows the application of silk layers to specifically patterned surfaces for sensing applications. Detecting variations at the nanoscale traditionally requires sophisticated instrumentation such as atomic force or electron microscopies, imaging based on dye-assisted spectroscopic techniques, or collective resonant effects in plasmonic structures, such as sub-wavelength apertures or surface enhanced Raman scattering. The precisely tunable thicknesses allow the convenience of interfacing protein films with resonant electromagnetic structures for enhanced detection approaches.

Two demonstrations of this capacity were demonstrated in the infrared (Altug, Adato et al. 2009) and optical regions of the electromagnetic spectrum (Omenetto, Lee et al. 2010) by spin coating silk protein monolayers on engineered nanoscale surfaces. In the first case, a silk monolayer was spin coated on plasmonic nanoantenna arrays to leverage the capacity to probe optical signatures from proteins in the mid-IR regime, where the vibrational fingerprint of the protein molecular structure was accessible. By tailoring the geometry of individual nanoantennas to form resonant structures that match the molecular vibrational mode there was enhancement at four orders of magnitude of the otherwise small IR signal. This approach allows measurement of the vibrational spectra from silk films at atto-mole levels because of the enhanced absorption signals from a small amount of molecules in the vicinity of the antenna nanotips. This enhanced response is further demonstrated by coating the nanoantennas with increasing (monolayer by monolayer) protein thicknesses.

In the optical regime, silk monolayers are spin coated on aperiodic photonic lattices consisting of the ordered arrangement of 200 nanometer diameter chromium nanoparticles on a quartz substrate. The silk protein monolayer was assessed for structural color in the visible range using conventional dark-field microscopy. This detection approach is label-free and relies on the confluence of photonics and biopolymer engineering by combining the nanoscale silk layers with nanopatterned aperiodic surfaces with controlled light scattering signatures.

The structure was sensitive to local changes in refractive index, giving rise to distinctive changes in the spatial and spectral color patterns observable under white light illumination (Plate XXI). When the slide is placed under a dark-field microscope, the white light from the condenser is scattered and spectrally rearranged into a structural color pattern. Additionally the multiple, deterministic spectral components encoded in the nano-quilt (angular spectra, scattering intensity, correlation patterns) yield distinct scattering responses which define a multi-parametric sensing platform for real-time nanoscale detection of biological materials in the visible spectral range.

Plate XXI (a) Nanopatterned aperiodic array of gold nanoparticles and its colorimetric response. The patterned surface occupies a 50 μim × 50 μim area. The inset shows a 5 μim × 5 μim area and the associated spectra extracted from a multispectral image of the lattice. By displaying only the red and green spectral challenge a colour transition is overtly observed (image b to image c). This transition corresponds to a change in topography of a silk monolayer, which is spin-coated on the structure. The atomic force microscope profiles below the images show the change in thickness corresponding to the different (and successive) spin-coating process.

Moreover, the recently demonstrated ability of the silk films to stabilize biochemical dopants as described earlier allows for expanded detection schemes exploiting the selectivity and responses of these biological components as active components in these engineered colorimetric chips. This approach could ultimately yield sensitivity limits comparable to surface plasmon sensors within a simpler, cost-effective approach with microfluidics, bio-assays, and label-free detection of biological materials on the nanoscale all accessible.

6.3.4 Bioresorbable electronics

These properties have been exploited towards devices that rely on ultrathin electronics supported by bioresorbable substrates of silk. When these devices were placed on living tissue, controlled hydration causes the silk to dissolve and resorb, initiating spontaneous, conformal, wrapping of tissue driven by capillary forces at the tissue interface. A combination of experimental and theoretical studies of the materials and underlying mechanics explored the transferring of a conformal electrode fabricated on the silk substrate (with an initial Young’s modulus of Esilk = 2.8 GPa), onto the surface of a living feline brain for EEG neural mapping experiments (Rogers, Kim et al. 2009; Kim, Viventi et al. 2010). These concepts provide capabilities for implantable or surgical devices that lie outside of wafer-based technologies or known forms of flexible electronics (Fig. 6.10). Further, as outlined earlier, the rate of silk film resorption or degradation can be tailored, from minutes to years, depending on the mode of processing.

Similar to the conformal electrode work described earlier, silk materials are suitable candidates for advanced implantable electronic devices that can interface with the body and establish conformal contact with the curvilinear surfaces of various organs. Achieving biocompatibility can be challenging, due to the complex nature of the biological response to many organic and inorganic materials. Electronic devices should be constructed out of materials that are soluble an/or biodegradable. Alternatively, a large fraction of the device can be designed to resorb, such that only small, safe, components remain, minimizing adverse biological responses. With this in mind, silk was used as the principal constituent of largely resorbable devices constituted of thin-film silicon electronics, to yield flexible devices largely resorbed in vivo (Rogers, Kim et al. 2009). The use of silicon provides high performance, good reliability, and robust operation. In this case, as described earler, silk is attractive compared to other biodegradable polymers such as poly(glycolic acid) (PGA), PLA and collagen, because of its robust mechanical properties, the ability to tailor the dissolution and/or biodegradation rates from hours to years, the generation of non-inflammatory amino acid degradation products, and the option to prepare the materials at ambient conditions to preserve sensitive electronic functions (Kaplan and Vepari 2007; Kaplan, Murphy et al. 2008).

6.3.5 Nanoparticle-doped silk

The use of silk doped with bioactive components can also be useful to add more latitude to the applications described above. As described earlier, the silk solution can be easily functionalized by mixing in suitable dopants to functionalize the devices, similar to what is done for silk-based drug delivery applications. Many instances of doping silks have been reported, from the inclusion of metals (Omenetto, Tao et al. 2010a), inorganic components such as silica (Kaplan, Foo et al. 2006) and hydroxyapatite (Yao, Liu et al. 2008), silk particles (Wang, Rajkhowa et al. 2010), fluorescent dyes (Mondia, Amsden et al. 2010; Han, Tansil et al. 2011), and drugs (Meinel, Hofmann et al. 2006; Boison, Wilz et al. 2008; Pritchard, Wilz et al. 2009), to other polymers for dynamic devices responsive to temperature (Kaplan, Gil et al. 2010).

Recently, silk films doped with gold nanoparticles (Au-NPs) have been used as light-activated heating elements that were interfaced with commercial thermo-electronic components to wirelessly power micro devices. In these experiments (Omenetto, Tao et al. 2010a) Au-NP doped silk films were interfaced to thermally available thermoelectric chips by casting the solution on the active area of the chip. Using a continuous wave (CW) green laser with an output power up to 450 mW/mm2 at 532 nm to match the absorption of the Au-NPs induces a temperature increase in the silk layer, which in turn is used to generate 20 mW of power. This use of silk solution underscores the ease of interfacing a doped protein with planar technologies. Power requirements of the illuminating source can be further optimized by either increasing the concentration of the Au-NPs and/or the thickness of the Au-NP doped silk film.

6.4 Conclusion

These outcomes are a promising prelude to opportunities that seamlessly combine technological function with biological function by leveraging the favorable processing environment and material features of silk proteins.

Silk fibroin films offer transparency, ability to nanopattern, and water-processing enabling the confluence of optical physics with biomaterials and the biomedical sciences. The ability to embed and maintain biological activity within a robust micro/nanostructured optical material is particularly unusual. Free-standing optical elements can be engineered to contain compounds across varying ‘biological scales’, from small organics, to complex proteins such as hemoglobin, and enzymes, while stored at room temperature.

Additionally, an important feature of these devices is their full biodegradability and biocompatibility, which expand the application space for silk-based devices in the environmental and life sciences where these attributes are paramount. The refined optical, electronic, and electro-optical functions obtainable through the various methods and approaches described herein would then be available for a class of new devices that could unobtrusively monitor and be part of a natural environment, including the human body. With this material versatility, devices for implantation in vivo become feasible, eliminating the need to retrieve the system at a later point. Such controlled degradation and biocompatibility would allow for silk-based technological devices to be dispersed in the environment as well, opening new opportunities for distributed sensing and disposable detection systems.

The availability of a free-standing biological matrix that has the material toughness to withstand room temperature use under ordinary environmental conditions while simultaneously exhibiting high technological quality and biological activity is unique. This platform could open opportunities for a new class of devices by weaving together the technological and biological worlds with a silk thread.

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