Nanobio-Approaches For The Detection Of Bacteria By Sers

Abstract

The main objective of this PhD thesis is to develop nanobio-based detection/identification protocols for pathogenic bacteria. Escherichia coli (E.coli) was the main target bacteria, Staphylococcus aureus (S.aureus) and Salmonella infantis (S.infantis) were included for comparison. Bacteria and their specific bacteriophages (used as specific bioprobes) were obtained from the expert laboratories and American Tissue Culture Collections (ATCC), and propagated, characterized and used by us. To enhance the optical signals, several plasmonic nanoparticles namely silver nanospheres (AgNPs), gold nanospheres and nanorods (AuNPs and AuNRs) were synthesized, characterized and the selected ones were used in the further studies described below. The SERS experiments were conducted in the following two different protocols: (i) The plasmonic nanoparticles were deposited on the carrier matrices, target bacteria were dropped and the SERS spectra were collected, then bacteriophages were added and changes in the spectral peaks were observed; (ii) the bacterial suspensions were mixed with the gold nanorod emulsions, then they were dropped on silica slides to collect the spectral data, then bacteriophages were added onto those surfaces and spectral changes with time were obtained. Especially the second protocol was very successful. The LSPR studies were performed on glass slides coated with polydopamine - carrying also the nanoparticles aggregates. We were able to follow a similar protocol that we applied in the SERS I experiments and changes in the LSPR spectra were used to describe the specific interaction of the target bacteria with their specific phages. Descriptive spectra for detection of three bacteria and iv their respective phages using the AuNPs were also obtained with a MALDI-TOF MS system. Especially changes in the specific phage peaks with time were found very descriptive. In the final part of the studies, the cross-linked gelatin microbeads were prepared and T4 phages were loaded within these gelatin hydrogel microspheres by a very simple technique. The results of loadings and release studies were concluded as the proof of concept of improving storage and release characteristics in use of bacteriophages effectively. Key words: Pathogenic bacteria, Detection, Bacteriophages, Nanoparticles, SERS, LSPR, MALDI-TOF MS, Gelatin hydrogel microspheres, Phage loading and release.