Bacteria capture using functionalized magnetic nanoparticles to facilitate the LC-ESI-MS analysis of foodborn pathogens
| Autor: | Yi-Ching Liu, 劉怡靜 |
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| Rok vydání: | 2008 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 96 Rapid identification of pathogens is of most importance considering the timeliness of therapy treatment. Traditional methods usually taking at least two days for pathogens incubation and detection fail to meet the aforementioned challenge. Recent progress in magnetic nanoparticles (MNPs) research has gained attention of their use for sample pretreatment and separation. These magnetic nano-materials could be modified with different kinds of functional group targeting to specific compounds and easily be isolated from matrix. Higher efficiencies and fast speed of analysis is also expected based on the property of high surface area per unit volume. This motivated us to couple these potential advantages to develop a technique of rapid identification for pathogens. MNPs prepared by chemical co-precipitation and modified with vancomycin were used as bacterial probes (MNPs-Van) which are specific to Gram-positive bacteria. Vancomycin was successfully immobilized onto the surface of MNPs with an average diameter of 10 nm as demonstrated by TEM, TGA and FTIR analysis. An aliquot of 200 mg bacterial probes (MNPs-Van) were applied to separate and trap Gram-positive bacteria (2×108 cfu mL-1)such as Staphylococcus aureus and Bacillus cereus from matrix with the assistance of external magnetic field (3000 G). After testing ultrasonication, freeze/thaw and microwave assisted cell lysis, we choose freeze/thaw method to extract the bacteria proteins. Specific biomarkers of proteins extracted from trapped bacteria were successfully identified by LC-ESI-MS analysis followed by database search. The detection of bacteria by bacterial probes (MNPs-Van) by LC-ESI-MS analysis and protein database search was successfully developed in this study. Bacteria could be identified based on individual protein molecular weight and retention time. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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