Manufacture of SU-8 Micro-grippers for Mechanical Characterization of Gut Epithelial Cells
This paper describes the manufacture of an integrated micro-electro-mechanical system (MEMS) to be used for small scale tissue manipulation. The micro-grippers are to be used to test the mechanical cell adhesion properties of the gut epithelium. In the majority of sporadic colon cancers the Adenomatous Polyopsis Coli (APC) protein is mutated or missing. Mutations of APC occur extremely early in the development of cancer, before formation of polyps. The following paper looks at the manufacturing processes for SU-8 micro-grippers. The micro-gripper structure has been successfully fabricated as a single structure not incorporating conduction paths. A number of sacrificial layers have been examined for compatibility with the SU-8 development. The sacrificial layer will be removed to leave a suspended structure. To avoid stiction between micro-grippers and the surface substrate the sacrificial layer must have a thickness of at least 6µm. An electroless coating process for Ni-P sacrificial layer has been developed. Coating Ni-P onto bare silicon substrates is a challenging problem. Sensitization and activation steps are needed to coat non-conductive surfaces. Sensitizers for Ni-P coatings of SnCl2 and 3-aminopropyltriethoxysilane were tested to look at adhesion of Ni-P to the substrate. Results have shown Ni-P can be applied to SiO2 surfaces at a thickness high enough to be utilized as a sacrificial layer for micro-grippers.
Micro-electro-mechanical systems, micro- gripping, biomechanics, SU-8, Nickel-phosphorus
The majority of sporadic colon cancers occur when the adenomatous polyopsis coli (APC) protein is mutated or lost . APC supports cell migration and cell adhesion . Loss of cell migration causes a build up of cells in a highly toxic environment. This causes mutations to occur within the cells. APC loss occurs very early in the progression of colon cancer and occurs before the formation of polyps . This project aims to characterize the mechanical properties of the gut epithelium cells and tissue, in particular looking at cell adhesion forces with the APC protein active or inactive.
Micro-gripping of constructs smaller than 50μm diameter is a challenging problem. Several types of micro-grippers have been reported. Shape Memory Alloy grippers can only be used for a limited number of cycles before the shape memory alloy fails or loses its shape memory effects . Piezoelectric actuators show high gripping forces with accurate displacement; however they need high operating voltages (10-100V) and require amplification methods to obtain large displacements .
Thermoelectric bimorph micro-grippers are of great interest. Luo et al  developed three types of micro-grippers, including bimorph grippers, whilst examining temperature rise and displacement. The results show small actuations of bimorph micro-grippers which operate in a small power range, however Type III micro-grippers, initially developed by Lin et al  with two horizontal hot arms yields largest displacement for given power input. This paper will develop the manufacturing techniques of the Type III tweezers to allow them to be used within tissue engineering conditions. Two materials will be investigated, Ni-P and SU-8. SU-8 is a non-conductive polymer however by adding a conductive metal circuit to the micro-grippers actuation is possible [8; 9]. Ni-P is a material currently utilized for biocompatible coatings of medical devices. The use of Ni-P as a bioMEMS material could simplify the fabrication process because it is conductive.
II. Design Concepts
A. Design Concept for the System
The design concept is shown in Figure 1. The structure will be manufactured using well established MEMS processes on a silicon chip. The structure includes a stage with incorporated micro-grippers for holding cell or tissue samples. A piezoelectric actuator will be used to stretch the tissues (which is attached to the right end of the tissue specimen). The deflection of the spring structure will be measured using an optical fiber sensor; critical forces can then be derived. Removal of the substrate under the micro-grippers will reduce stiction in biological environment and allow for smooth actuation.
B. Design of the Micro-Gripping System
A schematic of the micro-tweezers is shown in Figure 2. Design and finite element analysis was shown in previous publications [10; 11].
III. experimental procedures
SU-8 micro-grippers are to be manufactured using a three mask process (Fig. 2). Currently two types of SU-8 have been used for spin coating, SU-8 2007 and SU-8 2015 (Chestech Ltd., UK). SU-8 was spun onto 100 mm wafers with a thermally grown 200nm Si02 layer. Wafers were cleaned using acetone (VWR Internationl, UK) and then dried with N2; they were then rinsed with iso-propanol (IPA, VWR Internationl, UK) and dried with N2. Wafers were placed in an oven and heated to a temperature of 2000C; the heating step helped increase adhesion of SU-8 to SiO2. Spinning of SU-8 is done in three stages; the first stage helps spread the SU-8 at a low speed and acceleration. Stage 2 spin speed is selected according to the data sheet for a specific thickness. Stage 3 spin speed is higher still but for a short time period, around three seconds to help edge bead removal. The edge bead removal step is important to ensure the mask and wafer is in full contact for patterning. This results in a more precise structure being formed. After spinning the SU-8 resist, the sample is soft baked. Soft bake occurs for 1 min at 650C and 3 mins at 950C. A hotplate is used instead of an oven to ensure complete diffusion of solvent from the SU-8. The wafer is then placed in an OAI J500 mask aligner. The wafer height is raised to ensure full contact with the mask. The mask is placed chrome down to ensure highest resolution of the micro-grippers. The sample is then exposed to UV light with an intensity of 595mJ/cm2 for 30 seconds. A post exposure bake (PEB) is used to cure the SU-8 subjected to UV light; PEB parameters are equivalent to soft bake parameters. EC solvent (Chestech Ltd., UK) was used to develop structures; the sample was developed for i.e. 3 minutes. Finally the SU-8 structure was rinsed with IPA and dried with N2. SU-8 structures were examined using a Dektak stylus and optical microscope.
Microgripper fabrication: (A) Si wafer with 200nm layer SiO2 is coated with Ni-P (B). Ni-P is patterned and etched (C), the Cu/Cr electrode path is deposited on the sacrificial layer (D), this is patterned and etched (E). SU-8 is spin coated (F), this is patterned and developed (G). Finally the sacrificial layer is removed and Si etched to leave a suspended structure (H).
The final micro-gripper structure will incorporate suspended gripper arms (Fig. 3). To avoid stiction of the SU-8 micro-grippers to the Si substrate a sacrificial layer of high thickness (~6µm) is to be used. A number of materials were researched for the sacrificial layer. Initially Shipley S1818 was chosen as a sacrificial layer. Maximum thickness using S1818 was 2.5µm. The anchors were patterned using S1818; SU-8 was spun and developed on top of the patterned S1818. EC solvent is used to develop SU-8 structures; the EC solvent quickly removed the patterned S1818. S1818 was too thin a sacrificial layer and stripped easily in EC solvent which is utilized to develop SU-8. A sacrificial layer that would not etch in EC solvent or conduction path etching solutions was therefore needed. A possible solution was to use an electroless Ni-P coating.
Electroless Ni-P coatings can be coated using an autocatalytic process. Autocatalytic deposition can only occur on catalytic surfaces, e.g. Ni, Pd or steel. The SiO2 layer needs to be activated or coated with a catalytic material before Ni-P can be deposited. A number of groups have used SnCl2 (Sigma-Aldrich Company Ltd., UK) as a sensitizer and PdCl2 (Chestech Ltd., UK) as an activator [12; 13]. The wafers were cleaned using 10wt% NaOH (Sigma-Aldrich Company Ltd., UK); the solution was warmed to 500C to etch the top surface of the SiO2 layer. Wafers were rinsed with de-ionised water (DI water). Wafers were sensitized in tin chloride solution at 26.60C (70g/L SnCl2, 40g/L HCl). Wafers were rinsed again with DI water. Wafers were immersed in PdCl2 solution at room temperature (1g/L PdCl2, 10g/L HCl). Wafers were rinsed with DI water. Finally wafers were immersed in a Ni-P solution (Table I, Sigma-Aldrich Company Ltd., UK) at 880C with pH 4.8. The solution used deposits 10µm/hr. If wafers were activated successfully bubble formation occurs rapidly on the wafer surface followed by the solution turning black. Wafers were left in solution at 880C for 40mins.
The main challenge is to improve the adhesion of Ni-P to SiO2, a number of groups have used self assembled monolayers of 3-aminopropyltriethoxysilane (APTES, Sigma-Aldrich Company Ltd., UK) as a sensitizer layer to help improve adhesion [14; 15]. Wafers were cleaned in NaOH 10wt% solution at room temperature and rinsed with DI water. Then wafers were placed in a bath of NaOH 10wt% at 500C for ten minutes to etch the top surface of SiO2 followed by a bath of 90ml Toluene (Fisher Scientific UK Ltd., UK) and 10ml APTES for 2hrs at room temperature. Wafers were rinsed well with DI water before activation using PdCl2 solution; wafers were immersed in the solution for 30 seconds. Wafers were rinsed well with DI water and then placed in the Ni-P bath.
Table1Ni-P Bath Composition
Early results of SU-8 manufacture can be seen in figures 4 and 5. Structures were fabricated using SU-8 2015, structure of thickness up to 45µm have been successfully manufactured. Figure 5 shows the good resolution of the structures formed.
Sacrificial layers of Ni-P have been created up to a thickness of 20µm. Sensitization was found to be an important step. Immersion of wafers in SnCl2 solution followed by immersion in PdCl2 allowed SiO2 to be coated with Ni-P. Low adhesion between the wafer surface and Ni-P occurred resulting in the Ni-P layer peeling off the wafer during rinsing after removal from Ni-P bath. Sensitization using APTES followed by immersion in PdCl2 showed greater adhesion of Ni-P to SiO2 (Fig. 6). Ni-P layers were created up to 20µm in thickness. A small number of samples coated evenly and Ni-P had adhered well to the SiO2 substrate. Many samples, once removed from the Ni-P bath were unevenly coated with a very thin layer of Ni-P.
SU-8 2015 has been utilized to successfully create the micro-gripper structure. Finding an appropriate sacrificial layer that is compatible with the SU-8 processing steps has been a challenging problem. A novel Ni-P sacrificial layer which is biocompatible has been examined. Adhesion of Ni-P to SiO2 has been a problem; however use of self assembled monolayers has yielded some promising results. Other techniques such as sputtering a thin Ni layer (~100nm) and using this as the catalyst for electroless plating of Ni-P will be researched. Mechanical properties of Ni-P films will be examined.
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