Preparation Method And Instrument Investigation On Micropatterned Polymer Brushes With Selective Area Functionalization

With the rapid development of tissue engineering and regeneration medical, the interactions between cells and material interface have been attracting more and more academic interests. To artificially mimic the extra cell matrix (ECM) and investigate the cell-material interactions, surface patterning technique has been widely used in cell patterning, and presents immeasurable value in fundamental cell biology and high-throughput analysis research. Herein, in a interdisciplinary view of material science and processing equipment, the paper presents a practical study on three subjects:preparing Poly(HEMA) brushes functionalized graphene by surface-initiated activators regenerated by electron transfer atom transfer radical polymerization (SI-ARGET ATRP) and its biocompatibility assay,"Click" functionalization of micropatterned PEGEMAM brushes, and robot-aided microcontract printing system setup and its application in selective area functionalization of PEGMEMA micropatterns. The major innovations and results are as follows:(1) To explode the way to prepare high surface quality graphene nanosheets, comparative study on low temperature solvothermal reduction and hydrazine chemical reduction of graphene oxide was conducted. A method of low temperature solvothermal reduction of graphene oxide in NMP and GBL was proposed and characterized by ATR-FTIR, Raman, XPS, TGA, XRD, TEM, AFM, et al. In comparison, hydrazine reduction of laurylamine functionalized graphene oxide was also studied. Series characterizations indicated that hydrazine reduction resulted better surface quality graphene nanosheet without affecting the functionalization state of graphene surface. By SI-ARGET ATRP technique, the author succeeded to graft Poly(HEMA) and Ploy(HEMA-b-CA) brushes on graphene surfaces without Cu(II) precursor complex residual. The obtained functionalized graphene was sprayed onto glass slides. The BSA absorption and NIH-3T3cell culture experiments were performed on the obtained film scaffold, and results indicated that the amount of carboxyl groups contained in polymer brushes structure determined the BSA absorption capacity and NIH-3T3cell adhesion and proliferation behavior.(2) Combining microcontact printing and SI-ARGET ATRP, micropatterns of PEGMEMA brushes were prepared and testified it long-term anti-fouling property by cell culture experiments. A sequential terminal halogen substitution protocol was proposed to functionalize the PEGMEMA brushes:the terminal bromine of PEGMEMA brushes can be nucleophilic substituted by sodium azide, ethanolamine, and propargylamine, and finally reached PEGMEMA brushes with terminal functionalization of azide, hydroxyl and alkyne groups. A tunable surface functionalization density was obtained by controlling the substitution reaction time. In addition, the presence of both the azido and the acetylene groups on multicomponent PEGMEMA brushes enables in situ ligand immobilization via both the copper-free thiol-yne and strain induced Click chemistries within hNECs cell culture condition.(3) The challenges associated with precisely aligning and superimposing multiple μCP steps severely limits the extent of its applications. To overcome the issue, a vision guided selectively compliant articulated robotic arm was set up for investigating the feasibility of μCP applications (R-μCP). By image analysis using a custom MATLAB program, accuracy and precision of multiple-step R-μCP were obtained, which presented that the R-μCP system’s translational accuracy and precision in both the X and Y direction could be significantly reduced and consistently<15μm when aligning error compensation was programed into the SCARA’s alignment code. With the sequential R-μCP, SI-ARGET ATRP and PEGMEMA terminal substitution protocols, a precisely aligned tertiary (i.e., azido-, acetylene-, and non-functionalized) multicomponent brush composites was obtained. With the similar protocols, a precisely aligned tertiary multicomponent brush composites with gradient functionalization degree was also obtained, and further applied as cell culture substrate in hNECs culture experiments.