Fabrication of Cantilever and Fixed-fixed Beam Arrays (MEMS Devices). The fabrication protocol for these MEMS devices is as follows:
 Cleaning and Growth of Silicon Dioxide Layer:
A silicon wafer of 500-550 μm thickness (type P, dopant boron, orientation <100>, 100 mm diameter and, resistivity 0-100) was taken and cleaned well with 20 mL of piranha solution (H2SO4:H2O2 = 9:1) for 5 min. It was washed repeatedly with distilled water to remove metallic and organic contaminants from the surface and then a 1 μm layer of silicon dioxide was thermally grown over it by nano pyrogenic furnace.
(a) Dehydration and Spin Coat: A properly cleaned wafer was first dehydrated at 250 ºC for 10 min by keeping it over a hot plate to evaporate all of the surface moisture. Then it was spin coated with AZ5214E, photoresist (PR), of nearly 1.5 μm thickness at 6000 rpm for 40 seconds and baked to about 110 ºC for 2 min to evaporate the solvents in PR.
(b) Alignment for pattern transfer: A 3 inch DRIE lithomask was fixed over the treated wafer and kept under UV exposure for 2 sec in EVG 620 double sided mask aligner (50 mill joules/cm2). The mask was developed by dipping it in the solution of AZ351B:H2O in a ratio of 1:4 for 30 sec. It was washed with distilled water, dried under nitrogen and finally heated at 110 ºC for 4 min to get the pattern for the expected microstructures. The developed cantilevers and fixed-fixed beams were observed under the optical microscope.
 Dry Etch to release the pattern:
First formalin oil was put on a large carrier wafer to prepare sticky base for the sample. The sample was loaded inside the reactive ion etching chlorine (RIE-Cl) chamber and followed by the three steps.
(a) Anisotropic plasma etch of silicon: First silicon dioxide (SiO2) was plasma etched with 5 torr of pressure and 50 power for 6 min.
(b) Isotropic Si etch: Secondly silicon was etched with 7.5 torr of pressure and 30 power for 3 min and
(c) Oxygen etch: Lastly photoresist was removed by oxygen etching with 0 torr pressure and 150 power for 4 min.
The completely released devices were carefully diced from the wafer and proceeded for the functionalization steps. Four different cantilevers and fixed-fixed beams each of which distinguishable from the other based upon its length were fabricated. The cantilevers had the lengths of 86.6, 36.5, 28.1 and 19.8 μm respectively and the fixed-fixed beams had the lengths of 53.4, 35.5, 27.1 and 21.3 μm respectively. The cantilevers had the uniform width of 5.08 μm, whereas the fixed-fixed beams had uniform width of 4.74 μm. Each of the structure had a uniform thickness of 1.04 μm.
Covalent Surface Functionalization Protocols. The obtained silicon dioxide based microstructures were covalently functionalized by four different protocols (Figure 1). We had chosen B-doped silicon dioxide surface having free hydroxyl groups.
At first we have cleaned the surfaces of the silicon dioxide based substrates by dipping in 20 mL of piranha solution (H2SO4:H2O2 = 9:1) for 5 min to remove the organic and metallic contaminants. The temperature of the solution is maintained at 85 ºC and H2O2 was added carefully to replenish the temperature. These cleaned devices were undertaken for the further functionalization steps to create at least one anchor site with one or multiple amine groups.
Step 1: The hydroxyl groups on the cleaned surface of substrates were functionalized with 4% of 3-aminopropyl triethoxy silane (3-APTES) in 10 mL of toluene under N2 atmosphere at room temperature for 2 h. After the reaction time is over, the surfaces were rinsed with distilled water repeatedly and finally dried under nitrogen flow.
Step 1: The hydroxyl groups on the cleaned surface of substrates were functionalized with 10% of 3-mercaptopropyl triethoxy silane (3-MPTES) in 10 mL of toluene under N2 atmosphere at room temperature for 4 h. Then the surfaces were rinsed with fresh toluene repeatedly and finally dried under nitrogen flow. The surfaces were reacted immediately as described in the next step or it was kept under xylene to protect the surfaces from aerial oxidation.
Step 2: The free thiol groups on the surfaces were reacted with a crosslinker, 3-(maleimido)-propionic acid N-hydroxysuccinimide ester in 10 mL of dimethylformamide (DMF) for 5 h and then rinsed properly with water and ethanol. Again the surfaces were dried under nitrogen flow before proceeding for the next step.
Step 3: The surface immobilized N-hydroxysuccinimido group was further reacted with an amino functionalized polyhedral oligomeric silsesquioxane (POSS-NH2)16 moiety in 12 mL of DMF:H2O = 3:1 mixture. Ten equivalents of triethylmine (Et3N) and 0.1 equivalent of dimethylaminopyridine (DMAP) with respect to POSS-NH2 were also added to the same mixture and kept for overnight. It was rinsed properly with distilled water repeatedly and finally dried under nitrogen flow.
Step 1-3: These steps were similar as described in protocol 2.
Step 4: The aminated surface was then reacted with 10% of glutaraldehyde in 10 mL of PBS buffer (pH 7.4) for 4 h at room temperature. Then it was washed with distilled water and dried under nitrogen flow.
Step 5: This step is same as with the step 3 of protocol 2.
Step 1-5: These steps were similar as described in protocol 3.
Step 6-7: These steps are same as with the step 4 and 5 of protocol 3 respectively.
Optical Microscopy. The length and width of the MEMS devices were examined under Leica DIC optical microscope.
Scanning Electron Microscopy (SEM). The successfully released MEMS devices and the stability of them after all of the covalent functionalizations were monitored by Ultra 55, Field Emission Scanning Electron Microscope (FESEM) instruments from Carl Zeiss (Figure 2).
Atomic Force Microscopy (AFM). AFM images were obtained by JPK instruments using NanoWizard JPK00901 software in tapping mode. Analyses of the AFM images were processed using JPK data analyzer software. The cleaned surfaces were glued over a plate using a very small piece of double-sided tape and the images were recorded using silicon AFM tip with a resonance frequency about 300 kHz and force constant of 40 N/m.
X-ray Photoelectron Spectroscopy (XPS). In order to determine the surface abundance of atoms, X-ray photoelectron spectroscopy (XPS) was performed using a Kratos Axis ULTRA spectrometer (Shimadzu) outfitted with a non-monochromatic Al Kα X-ray source (105 W). Samples were electrically grounded for XPS measurements, and the binding energy scale was referenced to the Fermi level. Analyzer pass energy for wide (survey) sccm was 160 eV and for high resolution was 20 eV. The accelerating voltage was 105 kV and the current was 10 mA. Lense mode was kept in hybrid and SAC and STC vacuum level was kept at 1.8 e-8 and 1 e-7 torr respectively during the data recording. Samples were ultra-cleaned prior recording of the XPS data to remove the surface availability of unwanted atoms.
Time of Flight-Secondary Ion Mass Spectrometry (ToF-SIMS). To determine the nature of atomic and molecular species from the functionalized solid surface, we used time-of-flight secondary ion mass spectrometry (ToF-SIMS).17 This was performed using PHI TRIFT V nanoTOF model manufactured by Physical Electronics, USA. The data were acquired in the static mode using single beam in surface analytical regime where the samples were kept in UHV mode for surface acquisitions and sample chamber vacuum was typically 6.2e-7 Pascal region of pressure. Acquisition was done by a focussed 30KV Ga ion source in the LMIG gun which was rastered in 300 × 300 μm area to induce the desorption and ionization of atomic and molecular species from the functionalized surfaces with the beam current typically around 7 nA. The resulting secondary ions were accelerated into the mass spectrometer where they were mass separated by measuring the time-of-flight from the sample to the detector and a mass spectrum was recorded. Surface spectra were acquired in positive ion mode with a mass range of 0-1500 amu on 3-5 sections, for 5 min each, on two samples per set. A 2D Image was generated by rastering a finely focussed ion beam across the sample surface.
Laser Doppler Vibrometry (LDV). Advanced 3-D dynamic response data were collected from the functionalized MEMS devices by MSA-500 (Polytech) instruments before and after the attachment of the pheromones (Figure 3). The resonant frequency due to the base vibration was measured by the laser doppler vibrometer (LDV) to quantify the change in displacement and velocity of the vibrating structures without making any surface contact. The measured resonant frequency was extracted by analyzing the vibration spectrum with the use of polytech acquisition software.