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Self-Aligned Process for the Development of Surface Stress Capacitive Biosensor Arrays
Available online at www.sciencedirect.com
ProcediaProcedia Engineering 00 (2011) 000–000 Engineering 25 (2011) 835 – 838
Procedia Engineering www.elsevier.com/locate/procedia
Proc. Eurosensors XXV, September 4-7, 2011, Athens, Greece
Self-Aligned Process for the Development of Surface Stress Capacitive Biosensor Arrays Vasiliki Tsoutia*, Myrto Filippidoua, Christos Boutopoulosb, Panagiotis Broutasa, Ioanna Zergiotib, Stavros Chatzandroulisa a
Institute of Microelectronics, NCSR “Demokritos”, Terma Patriarchou Grigoriou, Agia Paraskevi 15310, Greece b Department of Applied Sciences, National Technical University of Athens, Zografou 15780, Greece
Abstract The fabrication process of a surface stress based capacitive biosensor array is presented. Flexible membranes and a fixed electrode on the substrate constitute the capacitive biosensors. Probe molecules are immobilized on the membrane surface and the surface stress variations during biological interactions force the membrane to deflect and effectively change the capacitance between the flexible membrane and the fixed substrate. The array consists of 60 sensors and thus is suitable for parallel sensing. Through the presented fabrication process, which is described in detail, sensitive membranes can be created. The process simplicity and the increased sensitivity of the biosensors are related to the use of boron implantation and the consequent self-aligned creation of the conductive membranes after the silicon fusion bonding technique.
© 2011 Published by Elsevier Ltd. Biosensor array; sensor fabrication; capacitive biosensors; DNA detection
1. Introduction Surface stress based biosensors utilize a flexible structure, usually a cantilever or a membrane. Their response depends on the surface stress changes induced during the interaction between the probe molecules immobilized on the flexible structure’s surface and the appropriate target molecules. They have attracted considerable interest as they detect biomolecular interactions through direct and simple methods.
* Corresponding author. Tel.: +30 210 650 3412; fax: +30 210 651 1723. E-mail address: [email protected].
1877-7058 © 2011 Published by Elsevier Ltd. doi:10.1016/j.proeng.2011.12.205
836 2
Vasilikiname Tsouti et al. / Procedia Engineering 25 000–000 (2011) 835 – 838 Author / Procedia Engineering 00 (2011)
Their main advantage is label-free sensing and consequently simplified sample preparation procedure and reduced cost compared to methods that involve labeling. In most cases, surface stress based biosensors are microcantilevers with optical or piezoresistive readout [1]. However, optical setups are difficult to implement, thus limiting their applications in practical systems. In addition, optical detection is difficult in opaque liquids. On the other hand, piezoresistive detection is temperature dependent and less sensitive. Capacitive detection is highly sensitive and requires low power consumption but is not feasible in cantilever biosensors due to faradic currents between the capacitor plates in an electrolyte solution. An alternative approach is to replace cantilevers with an ultrathin Si membrane effectively creating a sealed capacitor which prevents liquid insertion between its plates, thereby enabling reliable detection. Such structures hold promise for portable systems. In this work we present a new fabrication process for a biosensor array with sensing elements similar to those presented in [2-3]. There are two main differences in this fabrication process, which is simpler and less expensive than the previous one [2]. The first is that boron implantation (instead of an epitaxial SiGeB layer) is used for rendering the Si membrane conductive. The implantation results in rough sensor surface, which is expected to enhance the sensing performance of the membranes [4]. The second difference also renders the Si membrane more sensitive as the self-alignment of the membrane during boron implantation and the Silicon Fusion Bonding (SFB) technique allows for the creation of a narrow supporting rim. When the rim is narrower the membrane can deflect more effectively. 2. Biosensor Array Description and Fabrication Process The biosensor array consists of 60 membranes which are used as sensing elements for multiple target detection (up to 60). Each sensing element is a capacitor comprising two electrodes (Fig. 1a). The flexible electrode is a boron doped ultra thin silicon membrane suspended over a cavity. The counter electrode on the substrate is formed by phosphor implantation and is common for all sensing elements. The distance between the capacitor plates is determined by the thickness of the silicon dioxide layer that supports the membranes peripherally. The device operation relies on the surface stress changes induced on the membrane surface due to biomolecular interactions. In particular, probe molecules are immobilized on the membrane surface and waiting for binding with their respective target counterparts. The surface stress changes, mainly due to target-probe interactions, result in the membrane bending and therefore in a change of the capacitance between the flexible and the counter electrode. Due to the translation of the membrane deflection into a capacitance change, label-free sensing, miniaturization and ease of detection are enabled. For the fabrication of the biosensors we need two wafers, the membrane wafer (A) and the substrate wafer (B), which are then silicon fusion bonded in atmosphere environment without the need for alignment (Fig. 1a). First, a membrane support layer is formed after thermal oxidation for a 5000Å thick silicon dioxide layer. The circular cavities are defined utilizing optical lithography and through CHF 3 anisotropic dry etching in a plasma reactor the sensor cavities in the oxide are formed. Boron ion implantation for the creation of the capacitor’s flexible electrode on the membrane wafer follows. The ion implantation and annealing conditions determine the thickness of the final membranes and these conditions are selected based on simulation results and wet etching test experiments. The chosen conditions were 2*1016 ions/cm2 with energy 150keV, followed by thermal annealing at 1050ºC for 1h resulting in the formation of 0.8μm thick membranes. On the front side of the substrate wafer (B), the fixed electrode is created through phosphor implantation (1015 ions/cm2, 20keV, annealing at 1000℃ for 20min). Subsequently, a 200Å thick silicon dioxide film covers the fixed phosphor doped electrode in order to be used for the prevention of short circuit in case a membrane touches the substrate. After the preparation of the two wafers, the silicon
837 3
Vasiliki Tsouti et al. / Procedia Engineering (2011)000–000 835 – 838 Author name / Procedia Engineering 0025 (20111)
fusion bonding (SFB) takes place for the formation of the final Si wafer which will accommodate the capacitive micro-membrane arrays. The membrane wafer (A) is then lapped and the wafer thickness is reduced down to about 50µm. The rest of the wafer is etched with EDP (Ethylenediamine/ Pyrocatechol/ H2O) and the Si membranes are patterned as wet etching stops at the highly boron doped regions. A SEM image of the structure after the wet etching is shown in Fig. 1d.
(b)
(c)
(a)
(d)
Fig. 1. (a) Sensor array matrix fabrication process; (b) Part of the fabricated array; (c) 3-D image of a sensing element. It is observed that the surface is rough due to boron implantation; (d) SEM cross-section image of the sensing element. The cavity, the Si membrane, the supporting SiO2 and the narrow rim are indicated.
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Vasilikiname Tsouti et al. / Procedia Engineering 25 000–000 (2011) 835 – 838 Author / Procedia Engineering 00 (2011)
Furthermore, optical lithography and anisotropic dry etching of silicon oxide are used for the formation and the opening of the substrate contacts. Afterwards, a 5000Å thick Al layer is deposited and the desired Al areas are defined with optical lithography. The Al is removed from the non-protected with resist areas with wet etching and is annealed in forming gas environment (at 320ºC for 30min) for better Al adhesion on the silicon surface. In order to create a passivation layer, which can also serve as the probe molecules’ immobilization layer, a SiO2 film (Low Temperature Oxide, LTO) is deposited with chemical vapor deposition. For the removal of the LTO from the Al pads optical lithography is used, followed by wet etching with BHF for 1min and dry etching with SF 6 for 5min. Finally, a set of biosensor arrays of ultrathin Si membranes is fabricated, resulting in sensors with membrane diameter 150, 200 and 250μm. These arrays are cut in 12x12mm2 dies and their response is measured using a specially designed hybridization chamber which encloses only the sensing area (5x5mm2) of each array. In Fig. 1 the main steps of the fabrication process as well as images of the final structures are depicted. As expected, we observe that the surface of the membranes is rough owing to the implantation of boron (Fig. 1c) and the rim supporting the membrane is very narrow as it is created only by the lateral diffusion of boron and thus results in self-alignment (Fig. 1d). The fabricated devices were tested and the first experimental results using the beta-thalassemia CD19 mutation indicated that the sensors are able to detect the hybridization of DNA in 18nM concentration (results not shown). 3. Conclusions A new fabrication process for a biosensor array with Si membranes as sensing elements is demonstrated. The process is characterized by simplicity and lower cost obtained through the selfalignment of the membrane. In addition, the increased sensor surface roughness due to boron implantation is expected to enhance sensitivity. The functionality of the sensors has been proved for the detection of the beta-thalassemia CD19 hybridization. A more detailed analysis of the biosensor features and performance is in progress. Acknowledgements The research activities that led to these results, were co-financed by Hellenic Funds and by the European Regional Development Fund (ERDF) under the Hellenic National Strategic Reference Framework (NSRF) 2007-2013, according to Contract no. MICRO2-45 of the Project“Microelectronic Components for Lab-on-chip molecular analysis instruments for genetic and environmental application” within the Programme "Hellenic Technology Clusters in Microelectronics - Phase‐2 Aid Measure". References [1] Fritz J. Cantilever Biosensors. Analyst 2008; 133:855–63. [2] Tsouti V, Chatzandroulis S, Goustouridis D, Normand P, Tsoukalas D. Design and fabrication of a Si micromechanical capacitive array for DNA sensing. Microelectron. Eng. 2008;85:1359-61. [3] Tsouti V, Boutopoulos C, Andreakou P, Ioannou M, Zergioti I, Goustouridis D et al. Detection of DNA mutations using a capacitive micro-membrane array. Biosens. Bioelectron. 2010;26: 1588-92. [4] Lavrik NV, Tipple CA, Sepaniak MJ, Datskos PG. Gold Nano-Structures for Transduction of Biomolecular Interactions into Micrometer Scale Movements. Biomed. Microdevices 2001;3:1:35-44.
ProcediaProcedia Engineering 00 (2011) 000–000 Engineering 25 (2011) 835 – 838
Procedia Engineering www.elsevier.com/locate/procedia
Proc. Eurosensors XXV, September 4-7, 2011, Athens, Greece
Self-Aligned Process for the Development of Surface Stress Capacitive Biosensor Arrays Vasiliki Tsoutia*, Myrto Filippidoua, Christos Boutopoulosb, Panagiotis Broutasa, Ioanna Zergiotib, Stavros Chatzandroulisa a
Institute of Microelectronics, NCSR “Demokritos”, Terma Patriarchou Grigoriou, Agia Paraskevi 15310, Greece b Department of Applied Sciences, National Technical University of Athens, Zografou 15780, Greece
Abstract The fabrication process of a surface stress based capacitive biosensor array is presented. Flexible membranes and a fixed electrode on the substrate constitute the capacitive biosensors. Probe molecules are immobilized on the membrane surface and the surface stress variations during biological interactions force the membrane to deflect and effectively change the capacitance between the flexible membrane and the fixed substrate. The array consists of 60 sensors and thus is suitable for parallel sensing. Through the presented fabrication process, which is described in detail, sensitive membranes can be created. The process simplicity and the increased sensitivity of the biosensors are related to the use of boron implantation and the consequent self-aligned creation of the conductive membranes after the silicon fusion bonding technique.
© 2011 Published by Elsevier Ltd. Biosensor array; sensor fabrication; capacitive biosensors; DNA detection
1. Introduction Surface stress based biosensors utilize a flexible structure, usually a cantilever or a membrane. Their response depends on the surface stress changes induced during the interaction between the probe molecules immobilized on the flexible structure’s surface and the appropriate target molecules. They have attracted considerable interest as they detect biomolecular interactions through direct and simple methods.
* Corresponding author. Tel.: +30 210 650 3412; fax: +30 210 651 1723. E-mail address: [email protected].
1877-7058 © 2011 Published by Elsevier Ltd. doi:10.1016/j.proeng.2011.12.205
836 2
Vasilikiname Tsouti et al. / Procedia Engineering 25 000–000 (2011) 835 – 838 Author / Procedia Engineering 00 (2011)
Their main advantage is label-free sensing and consequently simplified sample preparation procedure and reduced cost compared to methods that involve labeling. In most cases, surface stress based biosensors are microcantilevers with optical or piezoresistive readout [1]. However, optical setups are difficult to implement, thus limiting their applications in practical systems. In addition, optical detection is difficult in opaque liquids. On the other hand, piezoresistive detection is temperature dependent and less sensitive. Capacitive detection is highly sensitive and requires low power consumption but is not feasible in cantilever biosensors due to faradic currents between the capacitor plates in an electrolyte solution. An alternative approach is to replace cantilevers with an ultrathin Si membrane effectively creating a sealed capacitor which prevents liquid insertion between its plates, thereby enabling reliable detection. Such structures hold promise for portable systems. In this work we present a new fabrication process for a biosensor array with sensing elements similar to those presented in [2-3]. There are two main differences in this fabrication process, which is simpler and less expensive than the previous one [2]. The first is that boron implantation (instead of an epitaxial SiGeB layer) is used for rendering the Si membrane conductive. The implantation results in rough sensor surface, which is expected to enhance the sensing performance of the membranes [4]. The second difference also renders the Si membrane more sensitive as the self-alignment of the membrane during boron implantation and the Silicon Fusion Bonding (SFB) technique allows for the creation of a narrow supporting rim. When the rim is narrower the membrane can deflect more effectively. 2. Biosensor Array Description and Fabrication Process The biosensor array consists of 60 membranes which are used as sensing elements for multiple target detection (up to 60). Each sensing element is a capacitor comprising two electrodes (Fig. 1a). The flexible electrode is a boron doped ultra thin silicon membrane suspended over a cavity. The counter electrode on the substrate is formed by phosphor implantation and is common for all sensing elements. The distance between the capacitor plates is determined by the thickness of the silicon dioxide layer that supports the membranes peripherally. The device operation relies on the surface stress changes induced on the membrane surface due to biomolecular interactions. In particular, probe molecules are immobilized on the membrane surface and waiting for binding with their respective target counterparts. The surface stress changes, mainly due to target-probe interactions, result in the membrane bending and therefore in a change of the capacitance between the flexible and the counter electrode. Due to the translation of the membrane deflection into a capacitance change, label-free sensing, miniaturization and ease of detection are enabled. For the fabrication of the biosensors we need two wafers, the membrane wafer (A) and the substrate wafer (B), which are then silicon fusion bonded in atmosphere environment without the need for alignment (Fig. 1a). First, a membrane support layer is formed after thermal oxidation for a 5000Å thick silicon dioxide layer. The circular cavities are defined utilizing optical lithography and through CHF 3 anisotropic dry etching in a plasma reactor the sensor cavities in the oxide are formed. Boron ion implantation for the creation of the capacitor’s flexible electrode on the membrane wafer follows. The ion implantation and annealing conditions determine the thickness of the final membranes and these conditions are selected based on simulation results and wet etching test experiments. The chosen conditions were 2*1016 ions/cm2 with energy 150keV, followed by thermal annealing at 1050ºC for 1h resulting in the formation of 0.8μm thick membranes. On the front side of the substrate wafer (B), the fixed electrode is created through phosphor implantation (1015 ions/cm2, 20keV, annealing at 1000℃ for 20min). Subsequently, a 200Å thick silicon dioxide film covers the fixed phosphor doped electrode in order to be used for the prevention of short circuit in case a membrane touches the substrate. After the preparation of the two wafers, the silicon
837 3
Vasiliki Tsouti et al. / Procedia Engineering (2011)000–000 835 – 838 Author name / Procedia Engineering 0025 (20111)
fusion bonding (SFB) takes place for the formation of the final Si wafer which will accommodate the capacitive micro-membrane arrays. The membrane wafer (A) is then lapped and the wafer thickness is reduced down to about 50µm. The rest of the wafer is etched with EDP (Ethylenediamine/ Pyrocatechol/ H2O) and the Si membranes are patterned as wet etching stops at the highly boron doped regions. A SEM image of the structure after the wet etching is shown in Fig. 1d.
(b)
(c)
(a)
(d)
Fig. 1. (a) Sensor array matrix fabrication process; (b) Part of the fabricated array; (c) 3-D image of a sensing element. It is observed that the surface is rough due to boron implantation; (d) SEM cross-section image of the sensing element. The cavity, the Si membrane, the supporting SiO2 and the narrow rim are indicated.
838 4
Vasilikiname Tsouti et al. / Procedia Engineering 25 000–000 (2011) 835 – 838 Author / Procedia Engineering 00 (2011)
Furthermore, optical lithography and anisotropic dry etching of silicon oxide are used for the formation and the opening of the substrate contacts. Afterwards, a 5000Å thick Al layer is deposited and the desired Al areas are defined with optical lithography. The Al is removed from the non-protected with resist areas with wet etching and is annealed in forming gas environment (at 320ºC for 30min) for better Al adhesion on the silicon surface. In order to create a passivation layer, which can also serve as the probe molecules’ immobilization layer, a SiO2 film (Low Temperature Oxide, LTO) is deposited with chemical vapor deposition. For the removal of the LTO from the Al pads optical lithography is used, followed by wet etching with BHF for 1min and dry etching with SF 6 for 5min. Finally, a set of biosensor arrays of ultrathin Si membranes is fabricated, resulting in sensors with membrane diameter 150, 200 and 250μm. These arrays are cut in 12x12mm2 dies and their response is measured using a specially designed hybridization chamber which encloses only the sensing area (5x5mm2) of each array. In Fig. 1 the main steps of the fabrication process as well as images of the final structures are depicted. As expected, we observe that the surface of the membranes is rough owing to the implantation of boron (Fig. 1c) and the rim supporting the membrane is very narrow as it is created only by the lateral diffusion of boron and thus results in self-alignment (Fig. 1d). The fabricated devices were tested and the first experimental results using the beta-thalassemia CD19 mutation indicated that the sensors are able to detect the hybridization of DNA in 18nM concentration (results not shown). 3. Conclusions A new fabrication process for a biosensor array with Si membranes as sensing elements is demonstrated. The process is characterized by simplicity and lower cost obtained through the selfalignment of the membrane. In addition, the increased sensor surface roughness due to boron implantation is expected to enhance sensitivity. The functionality of the sensors has been proved for the detection of the beta-thalassemia CD19 hybridization. A more detailed analysis of the biosensor features and performance is in progress. Acknowledgements The research activities that led to these results, were co-financed by Hellenic Funds and by the European Regional Development Fund (ERDF) under the Hellenic National Strategic Reference Framework (NSRF) 2007-2013, according to Contract no. MICRO2-45 of the Project“Microelectronic Components for Lab-on-chip molecular analysis instruments for genetic and environmental application” within the Programme "Hellenic Technology Clusters in Microelectronics - Phase‐2 Aid Measure". References [1] Fritz J. Cantilever Biosensors. Analyst 2008; 133:855–63. [2] Tsouti V, Chatzandroulis S, Goustouridis D, Normand P, Tsoukalas D. Design and fabrication of a Si micromechanical capacitive array for DNA sensing. Microelectron. Eng. 2008;85:1359-61. [3] Tsouti V, Boutopoulos C, Andreakou P, Ioannou M, Zergioti I, Goustouridis D et al. Detection of DNA mutations using a capacitive micro-membrane array. Biosens. Bioelectron. 2010;26: 1588-92. [4] Lavrik NV, Tipple CA, Sepaniak MJ, Datskos PG. Gold Nano-Structures for Transduction of Biomolecular Interactions into Micrometer Scale Movements. Biomed. Microdevices 2001;3:1:35-44.