Ordered Porous Film Prepared Via Microemulsion Method And Its Application In Biological Systems

Micropattern of biomaterials has attracted considerable attention due to their usein tissue engineering, cell investigation and biological detection. Various methodshave been developed for the fabrication of biomaterial patterns, top-down techniquesare the widely used ones. These methods are feasible, but the expensive equipmentand complex operation steps make them inconvenient for their application in manyareas. Self-assembly methods are more convenient compared to the top-downtechniques. Recently, breath figure method is attractive due to the use of cheap andnontoxic water droplets as template. However, the introduction of functional materialsalways realized by doping in organic solution or through complex introduction steps,which limit the functionalization with water soluble functional materials especiallywith brittle water soluble materials such as protein. Therefore, it is important todevelop a more simple method to achieve the fabrication of biomaterial pattern.In this dissertation, we develop a microemlsion method which also uses thecheap and nontoxic water droplets as template. We realize the convenient fabricationof biomaterial pattern through this microemulsion method. Using thehoneycomb-patterned porous film prepared by this microemulsion method as atemplate, ordered cell pattern can be obtained. Furthermore, the activities of cells onthe honeycomb-patterned porous film can be investigated. First, to simplify the introduction process of the water soluble materials, wedevelop a microemulsion method which also uses water droplets as template for thefabrication of ordered porous film. We use the microemulsion as casting solution,water droplets which come from the water phase of the microemulsion as template forthe honeycomb-patterned porous structure in this microemulsion method. Through thesize comparison of primary water droplets in micoemulsion and the pores in porousfilm, we found that the pores are much larger than the primary water droplets, thus thepores were formed by the fusion of primary water droplets in microemulsion.Furthermore, we also investigate the influence of solvent, polymer, surfactant, volumeratio of water phase and casting volume on the formation and order of the porous film.The surfactant distribute at oil-water interface to stabilize the water droplets due totheir amphiphilicity, thus the addition of surfactant is critical for the formation ofordered porous structure. The order and pore size of the porous film can be adjustedby varying the concentration of surfactant. In addition, solvent, polymer, volume ratioof water and casting volume can also influence the order and pore size of the porousfilm through affecting the evaporation speed of solvent and the stability of the waterdroplets. The investigations of these factors also confirm the existence of water fusionduring the formation of porous film. We propose a possible mechanism for thefabrication of ordered honeycomb-patterned porous film through microemulsionmethod based on the analysis of all the results, which is valuable for thefunctionalization of the porous film.Second, in the microemulsion method, the water droplets come from water phaseof the microemulsion, therefore, if we add the water soluble functional material intowater phase beforehand, it will be exist in the water droplets during the formation ofordered porous film and then fall down to the bottom of the pores with theevaporation of water, thus form the ordered porous film which was functionalized bythis water soluble material. Based on the above analysis, we dissolved the protein intowater phase aforehand, and then use this protein aqueous solution as a water phase toprepare the protein contained microemulsion. After casting this protein contained microemulsion onto proper substrate, protein contained porous film can be obtained.Through the confocal laser scanning microscopy characterization after proteinlabeling with fluorescent dye, we found that most of the protein locate at the bottomof the pores, thus form the similar pattern with the pores. Therefore, we can preparethe ordered porous film and protein pattern at the same time. Furthermore, thepatterned protein can interact with the molecular that can specifically recognize it,thus we can distinguish different kinds of protein using this protein pattern.Third, during the formation of ordered porous film, the surfactant locate at theoil-water interface, so it will be exist at the inner surface of the pores after the filmformation. In our system, we choose the positively charged DDAB as a surfactant,therefore, we can use this DDAB layer as a substrate for polyelectrolyte assemblybased on electrostatic interaction. After the assembly of positively chargedpolyelectrolyte on the inner surface of the pores, we can use this polyelectrolyte layeras a substrate to achieve the adhesion of negatively charged cell, and thus form thecell pattern similar to the array of the ordered pores. Here, we select the yeast cellswhich have the similar size to the patterned pores. Therefore, we can prepare orderedcell pattern with single cell in one pore, which is important for the investigation ofsingle cell. And the viability of the yeast cells retained through the FDA test.Furthermore, the yeast cells can grow larger till form the similar morphology with thepores after culturing for12hours, but their proliferation was inhibited due to the limitof the pores. The grown cells can proliferate again after releasing from the poresthrough immersing into sodium chloride solution, thus, we achieve the control of cellproliferation using our porous film, which is important for cell investigation, searchfor the genes which can control the cell size and shape, and the investigation ofdifferent kinds of diseases. In addition, if we assemble the negatively charged DNAand positively charged PEI on the inner surface of the pores before cell adhesion, wecan achieve the in-situ cell transformation in the pores. Moreover, the transformationefficiency increases with more deposition cycle.In a word, we develop a microemulsion method to achieve the fabrication of ordered honeycomb-patterned film. We can realize the convenient preparation of thewater soluble material pattern, such as protein pattern through this microemulsionmethod. In addition, through modifying the inner surface of the ordered pores, we canachieve the adhesion of cells in the pores, thus form ordered cell pattern. Cellinvestigations in the pores can also be taken based on this cell pattern. We believe thatthe results we got are very important for cell investigation, and the investigation andcontrol of many kinds of diseases.