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A fully biocompatible single-mode distributed feedback laser Cite this: Lab Chip, 2015, 15, 642 Received 4th October 2014, Accepted 20th November 2014

Yunkyoung Choi,a Heonsu Jeonab and Sunghwan Kim*c

DOI: 10.1039/c4lc01171f www.rsc.org/loc

A fully biocompatible laser would be attractive in many aspects of biomedical research. Here we report a single-mode biocompatible distributed feedback laser consisting of silk, riboflavin and silver in the form of a freestanding film. The distributed feedback structure has a large surface area and flexibility. The fabricated laser exhibited single-mode lasing at a wavelength of 495 nm.

Photonic-based bio/chemical sensors and diagnostic systems based upon measuring resonance-wavelength shifts of photonic resonators are attractive for characterizing biomolecular interactions.1–3 In particular, laser-based systems are ideal for biosensing and diagnostic purposes since their narrow resonance permits accurate resolving of small resonance shifts by low concentrations of biomolecules, along with the needlessness of high precision alignment for light-coupling and independence of sensitivity by biomolecular absorption.4 Numerous laser-based bio/chemical sensors have utilized the III–V compound semiconductor photonic crystals (PhCs),5,6 organic distributed feedback (DFB) lasers,7,8 and optofluidic ringresonators.9 In general, these devices are based on solid-state materials such as semiconductors, glasses, and polymers, for which biocompatibility remains an issue. An aim of current laser research for bioapplications is to demonstrate fully biocompatible lasers (FBLs), i.e. the active medium, matrix, and resonator are fully composed of biocompatible materials.10 The FBLs can be implanted into tissue and resorbed after use, thereby performing their function in vivo without limited penetration of light and contamination of the analytes. Since the pioneering studies investigating edible lasers in the early 1970s,11 gelatin and silk have been proved to be used as biocompatible laser matrices.12,13 And green a

Department of Physics and Astronomy, Seoul National University, Seoul 151-747, Republic of Korea b Department of Biophysics and Chemical Biology, Seoul National University, Seoul 151-747, Republic of Korea c Department of Physics & Department of Energy Systems Research, Ajou University, Suwon 443-749, Republic of Korea. E-mail: [email protected]

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fluorescent protein (GFP) and riboflavin (vitamin B2) have recently been adapted as biocompatible gain materials.12,14 Recently vitamin/glycerol microspheres encapsulated in patterned poly-L-lactic acid (PLLA) films have been reported as a completely biomaterial laser.15 However the laser resonator reported in that study resulted in multimode lasing action, but multimode spectra are undesirable for most applications, particularly biological applications. In contrast, organic DFB or PhC lasers offer lower thresholds and single-mode lasing properties. However the use of rigid substrates to support mechanically weak photonic resonators should be avoided for a single-mode FBL. In this study, we report, to the best of our knowledge, the first freestanding single-mode biocompatible DFB laser, with the entire laser structure composed of silk, riboflavin, and silver. The base material consisting of the periodic DFB structure and substrate was silk fibroin, a natural and biocompatible protein extracted from Bombyx mori cocoons. Riboflavin, better known as vitamin B2, was used here as a gain material that was doped into the silk DFB structure. A silver layer was inserted to optically isolate the active layer from the silk substrate. The surface corrugation on the active silk layer was transferred from the resist grating template generated via a laser-interference-lithography (LIL) method, an inexpensive and convenient fabrication method for sophisticated structures across large areas. Observations of single mode lasing at a wavelength of 495 nm and a threshold of 40 kW mm−2 were consistent with simulation results. A schematic of the fabrication procedure is illustrated in Fig. 1. First, we generated a photoresist (AZ MiR 701, MicroChemicals GmbH) grating with 250 nm thickness, 340 nm pitch size (Λ), and 30% duty cycle on silicon using the LIL method, as detailed in ref. 16. Scanning electron microscopy (SEM) images are shown in Fig. 2(a) and (b). Next, a blended aqueous solution of riboflavin and silk fibroin was prepared by mixing a 6% aqueous silk solution17 with a 10% riboflavin solution at a ratio IJv/v) of 1 : 1. The solution was then spincoated three times on the photoresist grating at a speed of

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Fig. 1 Schematic of the fabrication procedure and the fabricated biocompatible DFB laser.

2000 rpm (resulting thickness, ~750 nm). A silver isolate layer with 100 nm thickness was deposited by sputter-coating. For a biocompatible and flexible substrate, an aqueous solution of pure silk was poured on the deposited silver layer and allowed to solidify for 24 hours. The coagulated sample was immersed in acetone to selectively remove the photoresist grating, which detached the silk laser from the unnecessary silicon substrate without any deformation. We confirmed that the grating pattern was successfully decaled to the riboflavin-doped silk waveguide from the atomic microscopy (AFM) image (Fig. 2(c)) and the photographs showing good

Fig. 2 Scanning electron microscopy (SEM) images of (a) the top view and (b) the side view of the photoresist grating generated by the laserinterference-lithography (LIL) method. (c) Atomic force microscopy (AFM) image of the silk/riboflavin grating surface. (d) Green diffraction under white light. (e) Diffraction colors under bending of the fabricated DFB laser. All scale bars indicate 1 μm.

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diffraction even under bending of the FBL (Fig. 2(d) and (e)). The flexibility of our FBL offers potential for the development of human-implantable types of coherent light sources. Riboflavin (riboflavin 5′-monophosphate sodium salt hydrate, synthetic, molar mass: 455 g mol−1, CAS number 130-40-5 Sigma Aldrich) was selected as the gain medium for our FBL. This material is an oxidation–reduction catalyst that is compatible with the human body. Along with its biocompatibility, the beneficial optical properties of riboflavin were another reason that it was selected for our system. Riboflavin has a relatively high quantum yield of 0.23 and visible extinction/emission (near 450 nm/500 nm), while other biomolecules in the human body exhibit low quantum yields and must be excited in the ultraviolet range, which is biologically harmful.15 Notably, silk fibroin is well-suited to be a dispersing matrix of riboflavin for biological organic lasing. The aqueous and mild processing conditions to demonstrate the micro/nano-structures of silk fibroin have been shown to yield laser dye functions or the stable preservation of other biomolecules in silk films.18 In addition, in a previous study, silk fibroin was shown to present a lasing threshold one or two orders of magnitude lower than those in poly(methyl methacrylate) (PMMA) and DNA-CTMA when salt-derived laser dyes such as stilbene (or riboflavin used here) were doped since the repulsive electrostatic interaction between the dyes and the chains of silk fibroin might favourably affect the molecular arrangement of the dyes.13 Silver has been safely used in medicine for many years and is considered a biocompatible metal even though the abuse of silver might induce cytotoxic effects on the human body due to the release of silver ions.19 Basically all of the component materials of our FBL are biocompatible and the probable concern associated with the toxicity of the use of silver may be resolved by silk-encapsulation (top silk/riboflavin layer and bottom silk layer) and by using only trace amounts of silver.

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For optical characterization, the silk/riboflavin FBL was pumped with a frequency-tripled Nd:YAG laser of 5 ns duration, 10 Hz repetition rate, and a wavelength of 355 nm. The pumping beam was focused on the grating surface using a cylindrical lens. The focused beam spot size was 1 mm × 3 mm. The emitted light was detected using a spectrometer (Dongwoo Corp., DM700) along the normal to the grating surface. Fig. 3(a) shows the photoluminescence (PL) spectra of riboflavin in water and in the silk film. Since amino acids in silk such as tyrosine and tryptophan can stabilize the riboflavin complex, the riboflavin-doped silk film reveals a slightly red-shifted PL emission compared to that of the aqueous riboflavin solution.20 The emission spectra of the silk/riboflavin FBL are also shown in Fig. 3(b). For pumping above the lasing threshold, a narrow peak emerges at 495 nm with a full width at half maximum (FWHM) of 1.3 nm. The inset in Fig. 3(b) shows a plot of the output power as a function of pump energy. A

Fig. 3 (a) Photoluminescence spectra of riboflavin in water (blue) and in a silk film (green). The insets show photographs of the aqueous riboflavin solution and the riboflavin-doped silk film under excitation. (b) Measured spectra of the lasing emission at different pump intensities. The right inset denotes the output intensity as a function of the pump fluence. All scale bars indicate 1 μm.

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clear laser threshold behaviour is observed with a threshold of approximately 40 kW mm−2. This threshold value is rather high compared to thresholds obtained from organic DFB lasers using commercial laser dyes, which we attribute to the low quantum yield of riboflavin and the optical loss of the inserted silver layer. However, one highly promising aspect of our FBL is that the lasing threshold is of the same order of magnitude as that found in the reported metal-free riboflavin/gelatin DFB laser on the low refractive index grating,12 even though our FBL is accompanied by optical loss associated with the silver isolate layer.21,22 This result further demonstrates the favourable compatibility of the silk fibroin laser matrix with the biological riboflavin dye. Additionally it is noteworthy that our FBL exhibits stable output power under over 1 hour pumping exposure and for re-examination on the next day. This is because the high thermal conductivities of the silver layer and the silk layer with beta sheets23 enhance dissipation of heat generated by the pumping. The pitch size Λ of the fabricated surface grating was determined by λBragg = Λneff (second-order Bragg condition), where neff is the effective index of the waveguide optical mode.13 To investigate other photonic effects such as surface plasmons, numerical simulations were performed using the Lumerical software, based on the finite-difference timedomain (FDTD) method. In the simulations, the refractive index of the silk layers was 1.54. The complex dielectric constants of silver were taken from the Palik handbook.24 Fig. 4(a) shows the simulated spectrum. A strong and narrow peak was obtained at a wavelength of 497 nm, which is consistent with the experimental results. The normalized field intensity distribution of the magnetic field normal to the grating surface (TE polarization) which corresponds to the peak (Fig. 4(b)) shows that the surface grating induced the perturbation of the waveguide mode, thereby supplying

Fig. 4 (a) Simulated spectrum of the DFB structure. (b), (c) Normalized magnetic field (normal to the grating surface) intensity distributions corresponding to two resonance modes (λ = 497 nm for (b) and 465 nm for (c)).

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coherent feedback. We also investigated other resonance modes and showed a field intensity distribution corresponding to the peak at a wavelength of 465 nm in Fig. 4(c). These resonances originate from the same coherent feedback by the surface grating. But, higher order waveguide modes reduce the neff values of the silk/riboflavin DFB grating, thereby blue-shifting the coherent feedback resonances. Additionally we could not find any surface plasmon resonances which might affect or induce lasing behaviours for each of the polarizations due to the intrinsic loss in the metal.

Conclusions We demonstrated a fully biocompatible, single-mode operating, and free-standing second order DFB laser, assembled from a riboflavin-doped silk grating film, a silver optical isolate layer, and a silk substrate. The LIL method was used for generating the periodic surface grating across a large area. Single-mode lasing at 495 nm wavelength with 40 kW mm−2 was confirmed with photoluminescence measurements. In addition to their biocompatibility and optical transparency, silk films could be used as a good optical matrix material for organic lasers. Even though the silk/riboflavin active layer is considered to be in a solid state, hydrogel properties of the silk layer allow it to contain a large amount of water molecules and make larger channels for analytes in water to penetrate the silk layer. The silk film can preserve biological substances well such as enzymes and has good compatibility with cells. Along with the biocompatibility of our device, these favourable properties of silk promise fascinating potentials such as a highly sensitive biomolecular sensor and an optofluidic biolaser with sharp resonance peaks at which optofludic biolasers.25

Acknowledgements This work was supported by the Basic Science Program (NRF-2014R1A1A1008080) and the Nano-Material Technology Development Program (2009-0082580) through the National Research Foundation (NRF) and the NRF grant (2008-0061906) funded by the MSIP and MEST, Republic of Korea. It was also supported by the new faculty research fund of Ajou University.

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Notes and references 1 X. Fan, I. M. White, S. I. Shopova, H. Zhu, J. D. Suter and Y. Sun, Anal. Chim. Acta, 2008, 620, 8. 2 F. Vollmer and S. Arnold, Nat. Methods, 2008, 5, 591. 3 B. T. Cunningham and L. Laing, Expert Rev. Proteomics, 2006, 3, 271. 4 M. Zhang, C. Ge, M. Lu, Z. Zhang and B. T. Cunningham, Appl. Phys. Lett., 2013, 102, 213701. 5 M. Lončar, A. Scherer and Y. Qiu, Appl. Phys. Lett., 2003, 82, 4648. 6 S. Kim, J. Lee and H. Jeon, Appl. Phys. Lett., 2009, 94, 133503. 7 C. Vannahme, S. Klinkhammer, U. Lemmer and T. Mappes, Opt. Express, 2011, 19, 8179. 8 M. Liu, S. S. Choi, U. Irfan and B. T. Cunningham, Appl. Phys. Lett., 2008, 93, 111113. 9 X. Fan and I. White, Nat. Photonics, 2011, 5, 591. 10 M. C. Gather and S. H. Yun, Nat. Photonics, 2011, 5, 406. 11 T. W. Hänsch, Opt. Photonics News, 2005, 16, 14. 12 C. Vannahme, F. Maier-Flaig, U. Lemmer and A. Kristensen, Lab Chip, 2013, 13, 2675. 13 S. Toffanin, S. Kim, S. Cavallini, M. Natali, V. Benfenati, J. J. Amsden, D. L. Kaplan, R. Zamboni, M. Muccini and F. G. Omenetto, Appl. Phys. Lett., 2012, 101, 091110. 14 M. C. Gather and S. H. Yun, Nat. Photonics, 2011, 5, 406. 15 S. Nizamoglu, M. C. Gather and S. H. Yun, Adv. Mater., 2013, 25, 5943. 16 S. Ahn, S. Kim and H. Jeon, Appl. Phys. Lett., 2010, 96, 131101. 17 S. Kim, B. Marelli, M. A. Brenckle, A. N. Mitropoulos, E.-S. Gil, K. Tsioris, H. Tao, D. K. Kaplan and F. G. Omenetto, Nat. Nanotechnol., 2014, 9, 306. 18 E. M. Pritchard, P. B. Dennis, F. G. Omenetto, R. R. Naik and D. L. Kaplan, Biopolymers, 2012, 97, 479. 19 M. Bosetti, A. Massè, E. Tobin and M. Cannas, Biomaterials, 2002, 23, 887. 20 G. Blankenhorn, Eur. J. Biochem., 1978, 82, 155. 21 P. Andrew, G. A. Turnbull, I. D. W. Samuel and W. L. Barnes, Appl. Phys. Lett., 2002, 81, 954. 22 Y. M. Huang, F. Zhou and K. Xu, Appl. Phys. Lett., 2006, 88, 131112. 23 G. Liu, X. Huang, Y. Wang, Y.-Q. Zhang and X. Wang, Soft Matter, 2012, 8, 9792. 24 E. D. Palik, Handbook of Optical Constants of Solid, Academic, New York, NY, USA, 1985. 25 X. Fan and S. H. Yun, Nat. Methods, 2014, 11, 141.

Lab Chip, 2015, 15, 642–645 | 645

A fully biocompatible single-mode distributed feedback laser.

A fully biocompatible laser would be attractive in many aspects of biomedical research. Here we report a single-mode biocompatible distributed feedbac...
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