High-content protein microarray screening: Combining E. coli protein microarray with high-content screening to identify bacterial proteins that can influence neuronal morphogenesis
| Autor: | Lo, Peng-Lin, 羅鵬霖 |
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| Rok vydání: | 2015 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 103 Increasing evidence suggest that intestinal microbiota can influence the development and pathogenesis of the central nervous system. Therefore, in this study we attempted to examine the effect of each and every protein in E. coli proteome on neuronal morphogenesis. To identify all E. coli proteins that can alter neuronal morphology, we combined the protein microarray technique (which was originally developed to detect protein-protein interactions) with high-content screening (which was originally used to analyze cell morphology changes upon perturbation). To verify the feasibility of this method, we first established the culturing method to maintain mouse cortical neurons on the microarray surface. Second, we verified that protein spotted on microarray surface retained its activity using a known neurite outgrowth inhibitor. Third, we optimized the immunofluorescence staining protocol to maximize protein spot detect on this microarray. Finally, we used this novel platform to screen through the entire E. coli proteome and identify some proteins which can reduce neurite length or neuronal attachment. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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