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Advanced photonic biosensors for point-of-care diagnostics
Available online at www.sciencedirect.com
Procedia Procedia Engineering 00 (2011) Engineering 25000–000 (2011) 71 – 75
Procedia Engineering www.elsevier.com/locate/procedia
Proc. Eurosensors XXV, September 4-7, 2011, Athens, Greece
Advanced photonic biosensors for point-of-care diagnostics A.B. González-Guerreroa, S. Dantea, Daphné Duvala, J. Osmondb, Laura M. Lechugaa* a
Nanobiosensors and Bioanalytical Application Group, Research Center on Nanoscience and Nanotechnology (CIN2) CSIC& CIBER-BBN, Campus UAB, 08193 Bellaterra, Spain b Institute of Photonic Sciences (ICFO), 08860 Castelldefels, Spain
Abstract The application of portable, easy-to-use and highly sensitive biosensor devices for real-time diagnosis could offer significant advantages over current analytical methods. Silicon photonic biosensors based on evanescent wave detection have revealed themselves as the most promise candidates for achieving truly point-of-care devices. Advantages as miniaturization, extreme sensitivity, robustness, reliability, potential for multiplexing and mass production at low cost can be offered. We present our work towards the assembly of a point-of-care device by using integrated silicon interferometers sensors, polymer microfluidics and diffractive grating couplers for light incoupling. The micro/nanointerferometric devices have shown sensitivity close to 10-7 in refractive index units, which means an ability to discern, in a label-free scheme, concentrations of biological molecules at pM level.
© 2011 Published by Elsevier Ltd. Keywords: optical biosensor; lab-on-a-chip; interferometers; grating coupler.
1. Introduction Nomenclature LOC
Lab-on-a-chip
MZI
Mach-Zehnder interferometer
BiMW
Bimodal optical waveguides
DGC
Diffractive Grating Couplers
Most diagnostics techniques are based on time-consuming, expensive, and specialised techniques performed by trained technicians at laboratory level. These techniques typically require labelling of the samples or reagents with fluorescent or radioactive markers. It is clear that application of a portable, easy-
1877-7058 © 2011 Published by Elsevier Ltd. doi:10.1016/j.proeng.2011.12.018
722
A.B.L.González-Guerrero al. / Procedia 25 (2011) 71 – 75 Lechuga/ ProcediaetEngineering 00Engineering (2011) 000–000
to-use and highly sensitive lab-on-a-chip (LOC) platform for real-time diagnosis could offer significant advantages over current methods. Main applications fields of this technology will be clinical diagnostics, environmental monitoring, chemical and biological warfare surveillance, food industry and veterinary and industrial process control, among others. But despite remarkable progress towards LOC clinical assay systems, very few complete working prototypes have emerged [1]. Most of the LOC technology does not incorporate on-chip detection and the read-out must be done with complex scanners in laboratory settings. Incorporation of the “on-chip” detection by using biosensors is a new technology that shows great promise. Grating couplers
FUTURE OUTLOOK
Surface biofunctionalization Integrated Microfluidics
Optical readout
Array of Nanophotonic sensors
Electronics and data processing
Fig. 1. Scheme of our envisioned “lab-on-a-chip” device
Due to the scalable fabrication and the significant sensitivities (pM-fM) for biomolecular sensing, chipintegrated photonic array structures are excellent candidates as detectors for lab-on-a-chip applications (LOC). Integrated optical biosensors as grating-couplers, interferometers, photonic crystal, microring resonators or slot waveguides have been extensively studied in the last years [2-7] but few of them are commercially available yet. Interferometric biosensors have a broader dynamic range and show the best sensitivity levels achieved so far for label-free measurements (detection at the picomolar level, with variations of 10-7 in the refractive index-equivalent to 0.1pg/mm2) [6]. But the main drawback in the possible commercialization of the integrated interferometric devices is the complexity of the design, fabrication, optical adjustments and their apparently inherent instability, which is a consequence of their high sensitivity. In order to solve the above problems and for offering a reliable diagnostic tool, we are working in the development of LOC devices using interferometric micro/nanosensors based on silicon technology (see Figure 1). In the case of using integrated Mach-Zehnder Interferometer (MZI), a first Y-junction splits the guided light into two arms, the sensing arm and the reference arm which compensates for refractive index fluctuations and unspecific adsorption [6]. After a certain distance, the two signals are recombined into an output optical waveguide via a second Y-junction, producing the interference of both beams (Figure 1a). Moreover, MZI transducers present the possibility of developing biosensor microarrays, allowing the parallel analysis of a large amount of different samples on a single chip. Another innovative strategy which we have recently developed, is the bimodal waveguide (BiMW) interferometer [7], a single channel waveguide interferometer which mechanism is based on the interference of two waveguide modes of the same polarization (figure 1b). First, the light is coupled into a rib waveguide that supports a single transversal mode. After some distance, this mode is coupled into another waveguide which supports two transversal modes, the fundamental and the first order modes. Both modes are propagating until the output of the chip where they create an interference pattern. As the fundamental and first order modes have different intensity distribution at the core-cladding interface, the interference pattern is a function of the refractive index of the sensor area. The simplicity of the design of
A.B. González-Guerrero et al. / Procedia Engineering 25 000–000 (2011) 71 – 75 Author name / Procedia Engineering 00 (20111)
these bimodal waveguide (BiMW) interferometers is attractive for mass-fabrication as there is no further need for using Y-shape splitters, which are the most complex component of the MZI devices.
Fig 2. Scheme of a) a MZI and b) a BiMW interferometer
2. Implementation of the lab-on-a-chip The lab-on-a-chip is being assembled by integrating the following parts: (i) micro/nano sensors fabricated with standard silicon technology, (ii) a polymer microfluidic and the flow delivery system (iii) diffractive grating coupler for the in coupling of the light in all the photonic channels for multisensing (iv) reliable immobilisation and regeneration protocols for the biological receptor (iv) photodetectors, electronic & software control (v) final integration and packaging. 2.1. The sensor chip and the microfñuidics We employ dielectric waveguides (Si3N4) as core waveguides for both the MZI and the BiMW devices. Before fabrication, modeling and numerical simulation was done using Finite Element Beam Propagation methods for defining the proper parameters of the waveguides and layout with optimised power amplitude. The final layout design included dozens of chip-integrated waveguides into a single wafer substrate using silicon waveguide technology (Si/SiO2/Si3N4/SiO2). For the MZI, the waveguide core is 200 nm of Si3N4 (n=2.00). For the BiMW, the single mode part is 150 nm of Si3N4 (n=2.00) thick and the bimodal part is 350 nm thick. The rid height is only 2 nm for both devices. Both devices are designed for operation at visible wavelengths with monochromatic and polarized (TE or TM) light.
Fig. 3. (a) Picture of one chip with MZI devices; (b) Picture of one chip with 16 BiMW devices and sealed with the polymer microfluidics.
The flow system consists of a flow cell with four independent channels fabricated in PDMS. The integrated microfluidic/sensor platform is supplied with tube connections to the exterior. The flow delivery system includes a syringe pump and injection valves. Fig. 3b) shows a photo of one BiMW chip inside the flow cell.
73 3
74 4
A.B. González-Guerrero et al. / Procedia 00 Engineering 25 (2011) 71 – 75 L. Lechuga/ Procedia Engineering (2011) 000–000
2.2 Light incoupling Diffractive grating couplers allow an efficient light coupling into the waveguides with high coupling efficiency. Due to the dimensions of the interferometers and to the operating wavelength (visible range), the diffractive grating length cannot exceed 100 µm with a sub-micronic period. The grating has been directly written onto the Si3N4 waveguide by electron-beam lithography. Fig. 4 shows the layout of a MXI chip and the position of the grating. The grating period is 400 nm, with a duty cycle of 0.5 and a depth of 40 nm (total dimensions: 20 µm x 100 µm). Fig. 4 also shows photos of the grating couplers and light incoupling into a MZI device. 3.2 cm D0 W0 I1 D1 W1 I2 I3 I4 I5
1 cm W0 D0 W1 D1 W2 I2 I3 I4
grating
Rib WG
I5
Fig. 4. Layout of a MZI chip. Photos of the grating couplers and in-coupling of the light in a MZI device.
3. Sensitivity and biosensing evaluation The sensitivity of MZI and BiMW devices was checked by injecting decreasing concentrations of HCl. Recording phase variation versus refractive index variation for each concentration gives a detection limit of nmin=1.1×10-7 RIU and nmin=2.5×10-7 RIU for MZI and BiMW, respectively. These values indicate that the interferometric devices are one of the most sensitive sensors and could allow anultrasensitivity label-free detection of biomolecular interactions. For biosensing evaluation, proteins, bacteria and DNA have been tested and the devices have shown an excellent performance for label-free detection with the immunosensing of hormones at 1 pM level and bacteria detection at 10 cfu/ml level. 4. Conclusion We are working on the realization of a sensitive, affordable, hand-held and portable device for point of care diagnosis. The lab-on-chip device includes ultrasensitive interferometric biosensors based on integrated silicon-based optical waveguides which have already shown their ability to detect minute variation of the refractive index (Δn min =1.10-7) and label-free biosensing detection at pM level. Light in coupling into the sensor waveguides is achieved by diffraction gratings couplers fabricated by electron beam on top of the rib waveguides. Hermetic sealed microfluidics is achieved by using a polymer cartridge. Multiplexed excitation and read-out of all the sensors within a single chip is in progress. Acknowledgements Authors acknowledge the financial support from M. Botín Foundation, SENA project (TSI-020301-200811) from the Spanish Ministry of Industry and LEUKEM-CHIP project from CIBER-BBN. This work was partially developed using the micro and nano fabrication capabilities of the ICTS/clean room from the IMB-CNM (CSIC) under GISERV
A.B. González-Guerrero et al. / Procedia Engineering 25 (2011) 71 – 75 Author name / Procedia Engineering 00 (20111) 000–000
References [1] Ligler, F. S. Perspective on Optical Biosensors and Integrated Sensor Systems. Anal. Chem. 2008; 81:519–526. [2] X. Fan, I. M. White, S. I. Shopoua, H. Zhu, J. D. Suter and Y. Sun, Sensitive optical biosensors for unlabeled targets: A review . Anal. Chim. Acta, 2008; 620:8–26. [3] D. Erickson, S. Mandal, A. H. J. Yang and B. Cordovez, Nanobiosensors: optofluidic, electrical and mechanical approaches to biomolecular detection at the nanoscale. Microfluid. Nanofluid., 2008;4: 33–52. [4] F. Vollmer and S. Arnold,. Whisperry gallery mode biosensing: label-free detection down to single molecules. Nat. Methods, 2008; 5:591–596. [5] A.L. Wasburn and R.C. Bailey. Photonics-on-a-chip: recent advances in integrated waveguides as enabling detection elements for real-world, lab-on-a-chip biosensing applications. Analyist. 2011; 136: 227-236. [6] M.C. Estevez, M. Álvarez and L. M. Lechuga. Integrated Optical devices for lab-on-a-chip biosensing applications. Laser & Photonics Reviews, 2011 (In Press) [7] K. E. Zinoviev, A.B. González-Guerrero, C. Domínguez and L. M. Lechuga. Integrated Bimodal Waveguide Interferometric Biosensor for Label-free Analysis. J. Lightwave Tech. 2011; 29 (13): 1926-1930.
755
Procedia Procedia Engineering 00 (2011) Engineering 25000–000 (2011) 71 – 75
Procedia Engineering www.elsevier.com/locate/procedia
Proc. Eurosensors XXV, September 4-7, 2011, Athens, Greece
Advanced photonic biosensors for point-of-care diagnostics A.B. González-Guerreroa, S. Dantea, Daphné Duvala, J. Osmondb, Laura M. Lechugaa* a
Nanobiosensors and Bioanalytical Application Group, Research Center on Nanoscience and Nanotechnology (CIN2) CSIC& CIBER-BBN, Campus UAB, 08193 Bellaterra, Spain b Institute of Photonic Sciences (ICFO), 08860 Castelldefels, Spain
Abstract The application of portable, easy-to-use and highly sensitive biosensor devices for real-time diagnosis could offer significant advantages over current analytical methods. Silicon photonic biosensors based on evanescent wave detection have revealed themselves as the most promise candidates for achieving truly point-of-care devices. Advantages as miniaturization, extreme sensitivity, robustness, reliability, potential for multiplexing and mass production at low cost can be offered. We present our work towards the assembly of a point-of-care device by using integrated silicon interferometers sensors, polymer microfluidics and diffractive grating couplers for light incoupling. The micro/nanointerferometric devices have shown sensitivity close to 10-7 in refractive index units, which means an ability to discern, in a label-free scheme, concentrations of biological molecules at pM level.
© 2011 Published by Elsevier Ltd. Keywords: optical biosensor; lab-on-a-chip; interferometers; grating coupler.
1. Introduction Nomenclature LOC
Lab-on-a-chip
MZI
Mach-Zehnder interferometer
BiMW
Bimodal optical waveguides
DGC
Diffractive Grating Couplers
Most diagnostics techniques are based on time-consuming, expensive, and specialised techniques performed by trained technicians at laboratory level. These techniques typically require labelling of the samples or reagents with fluorescent or radioactive markers. It is clear that application of a portable, easy-
1877-7058 © 2011 Published by Elsevier Ltd. doi:10.1016/j.proeng.2011.12.018
722
A.B.L.González-Guerrero al. / Procedia 25 (2011) 71 – 75 Lechuga/ ProcediaetEngineering 00Engineering (2011) 000–000
to-use and highly sensitive lab-on-a-chip (LOC) platform for real-time diagnosis could offer significant advantages over current methods. Main applications fields of this technology will be clinical diagnostics, environmental monitoring, chemical and biological warfare surveillance, food industry and veterinary and industrial process control, among others. But despite remarkable progress towards LOC clinical assay systems, very few complete working prototypes have emerged [1]. Most of the LOC technology does not incorporate on-chip detection and the read-out must be done with complex scanners in laboratory settings. Incorporation of the “on-chip” detection by using biosensors is a new technology that shows great promise. Grating couplers
FUTURE OUTLOOK
Surface biofunctionalization Integrated Microfluidics
Optical readout
Array of Nanophotonic sensors
Electronics and data processing
Fig. 1. Scheme of our envisioned “lab-on-a-chip” device
Due to the scalable fabrication and the significant sensitivities (pM-fM) for biomolecular sensing, chipintegrated photonic array structures are excellent candidates as detectors for lab-on-a-chip applications (LOC). Integrated optical biosensors as grating-couplers, interferometers, photonic crystal, microring resonators or slot waveguides have been extensively studied in the last years [2-7] but few of them are commercially available yet. Interferometric biosensors have a broader dynamic range and show the best sensitivity levels achieved so far for label-free measurements (detection at the picomolar level, with variations of 10-7 in the refractive index-equivalent to 0.1pg/mm2) [6]. But the main drawback in the possible commercialization of the integrated interferometric devices is the complexity of the design, fabrication, optical adjustments and their apparently inherent instability, which is a consequence of their high sensitivity. In order to solve the above problems and for offering a reliable diagnostic tool, we are working in the development of LOC devices using interferometric micro/nanosensors based on silicon technology (see Figure 1). In the case of using integrated Mach-Zehnder Interferometer (MZI), a first Y-junction splits the guided light into two arms, the sensing arm and the reference arm which compensates for refractive index fluctuations and unspecific adsorption [6]. After a certain distance, the two signals are recombined into an output optical waveguide via a second Y-junction, producing the interference of both beams (Figure 1a). Moreover, MZI transducers present the possibility of developing biosensor microarrays, allowing the parallel analysis of a large amount of different samples on a single chip. Another innovative strategy which we have recently developed, is the bimodal waveguide (BiMW) interferometer [7], a single channel waveguide interferometer which mechanism is based on the interference of two waveguide modes of the same polarization (figure 1b). First, the light is coupled into a rib waveguide that supports a single transversal mode. After some distance, this mode is coupled into another waveguide which supports two transversal modes, the fundamental and the first order modes. Both modes are propagating until the output of the chip where they create an interference pattern. As the fundamental and first order modes have different intensity distribution at the core-cladding interface, the interference pattern is a function of the refractive index of the sensor area. The simplicity of the design of
A.B. González-Guerrero et al. / Procedia Engineering 25 000–000 (2011) 71 – 75 Author name / Procedia Engineering 00 (20111)
these bimodal waveguide (BiMW) interferometers is attractive for mass-fabrication as there is no further need for using Y-shape splitters, which are the most complex component of the MZI devices.
Fig 2. Scheme of a) a MZI and b) a BiMW interferometer
2. Implementation of the lab-on-a-chip The lab-on-a-chip is being assembled by integrating the following parts: (i) micro/nano sensors fabricated with standard silicon technology, (ii) a polymer microfluidic and the flow delivery system (iii) diffractive grating coupler for the in coupling of the light in all the photonic channels for multisensing (iv) reliable immobilisation and regeneration protocols for the biological receptor (iv) photodetectors, electronic & software control (v) final integration and packaging. 2.1. The sensor chip and the microfñuidics We employ dielectric waveguides (Si3N4) as core waveguides for both the MZI and the BiMW devices. Before fabrication, modeling and numerical simulation was done using Finite Element Beam Propagation methods for defining the proper parameters of the waveguides and layout with optimised power amplitude. The final layout design included dozens of chip-integrated waveguides into a single wafer substrate using silicon waveguide technology (Si/SiO2/Si3N4/SiO2). For the MZI, the waveguide core is 200 nm of Si3N4 (n=2.00). For the BiMW, the single mode part is 150 nm of Si3N4 (n=2.00) thick and the bimodal part is 350 nm thick. The rid height is only 2 nm for both devices. Both devices are designed for operation at visible wavelengths with monochromatic and polarized (TE or TM) light.
Fig. 3. (a) Picture of one chip with MZI devices; (b) Picture of one chip with 16 BiMW devices and sealed with the polymer microfluidics.
The flow system consists of a flow cell with four independent channels fabricated in PDMS. The integrated microfluidic/sensor platform is supplied with tube connections to the exterior. The flow delivery system includes a syringe pump and injection valves. Fig. 3b) shows a photo of one BiMW chip inside the flow cell.
73 3
74 4
A.B. González-Guerrero et al. / Procedia 00 Engineering 25 (2011) 71 – 75 L. Lechuga/ Procedia Engineering (2011) 000–000
2.2 Light incoupling Diffractive grating couplers allow an efficient light coupling into the waveguides with high coupling efficiency. Due to the dimensions of the interferometers and to the operating wavelength (visible range), the diffractive grating length cannot exceed 100 µm with a sub-micronic period. The grating has been directly written onto the Si3N4 waveguide by electron-beam lithography. Fig. 4 shows the layout of a MXI chip and the position of the grating. The grating period is 400 nm, with a duty cycle of 0.5 and a depth of 40 nm (total dimensions: 20 µm x 100 µm). Fig. 4 also shows photos of the grating couplers and light incoupling into a MZI device. 3.2 cm D0 W0 I1 D1 W1 I2 I3 I4 I5
1 cm W0 D0 W1 D1 W2 I2 I3 I4
grating
Rib WG
I5
Fig. 4. Layout of a MZI chip. Photos of the grating couplers and in-coupling of the light in a MZI device.
3. Sensitivity and biosensing evaluation The sensitivity of MZI and BiMW devices was checked by injecting decreasing concentrations of HCl. Recording phase variation versus refractive index variation for each concentration gives a detection limit of nmin=1.1×10-7 RIU and nmin=2.5×10-7 RIU for MZI and BiMW, respectively. These values indicate that the interferometric devices are one of the most sensitive sensors and could allow anultrasensitivity label-free detection of biomolecular interactions. For biosensing evaluation, proteins, bacteria and DNA have been tested and the devices have shown an excellent performance for label-free detection with the immunosensing of hormones at 1 pM level and bacteria detection at 10 cfu/ml level. 4. Conclusion We are working on the realization of a sensitive, affordable, hand-held and portable device for point of care diagnosis. The lab-on-chip device includes ultrasensitive interferometric biosensors based on integrated silicon-based optical waveguides which have already shown their ability to detect minute variation of the refractive index (Δn min =1.10-7) and label-free biosensing detection at pM level. Light in coupling into the sensor waveguides is achieved by diffraction gratings couplers fabricated by electron beam on top of the rib waveguides. Hermetic sealed microfluidics is achieved by using a polymer cartridge. Multiplexed excitation and read-out of all the sensors within a single chip is in progress. Acknowledgements Authors acknowledge the financial support from M. Botín Foundation, SENA project (TSI-020301-200811) from the Spanish Ministry of Industry and LEUKEM-CHIP project from CIBER-BBN. This work was partially developed using the micro and nano fabrication capabilities of the ICTS/clean room from the IMB-CNM (CSIC) under GISERV
A.B. González-Guerrero et al. / Procedia Engineering 25 (2011) 71 – 75 Author name / Procedia Engineering 00 (20111) 000–000
References [1] Ligler, F. S. Perspective on Optical Biosensors and Integrated Sensor Systems. Anal. Chem. 2008; 81:519–526. [2] X. Fan, I. M. White, S. I. Shopoua, H. Zhu, J. D. Suter and Y. Sun, Sensitive optical biosensors for unlabeled targets: A review . Anal. Chim. Acta, 2008; 620:8–26. [3] D. Erickson, S. Mandal, A. H. J. Yang and B. Cordovez, Nanobiosensors: optofluidic, electrical and mechanical approaches to biomolecular detection at the nanoscale. Microfluid. Nanofluid., 2008;4: 33–52. [4] F. Vollmer and S. Arnold,. Whisperry gallery mode biosensing: label-free detection down to single molecules. Nat. Methods, 2008; 5:591–596. [5] A.L. Wasburn and R.C. Bailey. Photonics-on-a-chip: recent advances in integrated waveguides as enabling detection elements for real-world, lab-on-a-chip biosensing applications. Analyist. 2011; 136: 227-236. [6] M.C. Estevez, M. Álvarez and L. M. Lechuga. Integrated Optical devices for lab-on-a-chip biosensing applications. Laser & Photonics Reviews, 2011 (In Press) [7] K. E. Zinoviev, A.B. González-Guerrero, C. Domínguez and L. M. Lechuga. Integrated Bimodal Waveguide Interferometric Biosensor for Label-free Analysis. J. Lightwave Tech. 2011; 29 (13): 1926-1930.
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