Preparation and surface-functionalization of dye-doped bioconjugated poly(styrene-co-maleimide) nanoparticles
Thesis (MSc)--Stellenbosch University, 2014.
ENGLISH ABSTRACT: Standard methods used for detection of microbial contamination in water and food are based on culturing of samples to increase the number of bacteria present to a detectable level. These tests are however time-consuming (requiring one to two days for results to be generated) and labour intensive. The study described in this dissertation aims to develop dye-doped poly(styrene-co-maleimide) nanoparticles conjugated to antibodies for the rapid (within a few hours) detection of E. coli in water via clustering of the nanoparticles around bacteria to create a concentrated and localised fluorescent signal. Pristine and dye-doped nanoparticles were synthesised according to a one-step (published) method, and surface-functionalised with formaldehyde and three different diamine compounds. The percentage dye added to the particles was also varied in the study. The functionalised nanoparticles were characterised via FTIR, TEM, DSC, and Zetasizer. The emission properties of the dye-loaded NPs were measured with a spectrofluorometer. The functionalised nanoparticles were bioconjugated with avidin and antibodies. The extent of avidin attachment was quantified via the use of a biotin(5-fluorescein) conjugate and via an avidin titration assay. The functionality of the bioconjugated nanoparticles was tested via a sandwich assay and a centrifuge based assay with E. coli cultures. Fluorescence microscopy was also utilised to demonstrate the utility of the particles in creating localised fluorescent signals indicating the presence of E. coli cells.