Intravascular catheters (IVCs) are one of the most frequently used medical devices in hospitals. However, catheters are associated with serious IVC-related bloodstream infection (IVC-BSI), which causes high mortality and additional costs. IVC provides surfaces for microbes attach to, grow and form a biofilm with the subsequent shedding or dispersion of microbes leading to bacteraemia. The formation of a biofilm by microbes is usually associated with IVC-BSI infection and often results in increased clinical pathology. Biofilms are highly resistant to a number of antimicrobial agents as well as the host immune system. While many studies have described biofilms, there are still deficits in our understanding of the mechanism of biofilm formation and multifactorial antimicrobial resistance. Therefore, it is urgent to improve our understanding on microbial biofilm on IVCs. This thesis is the first comprehensive study of Staphylococcus aureus (S. aureus) isolated from Queensland hospitals that assessed the biofilm characteristics of nine S. aureus isolates in terms of their capacity to form biofilms on commercial catheter surfaces, resistance against ampicillin and underpinning genetic components. The ability of S. aureus isolates to form a biofilm on microtiter plates or on two commercial catheter tips, Introcan-Safety® and SurFlash® was determined by Crystal Violet Assay (CV) or estimating CFU of biofilm cells using Catheter Tips Assay (CTA). S. aureus biofilm resistance was determined based on minimum inhibitory concentrations (MIC) and biofilmMIC (BMIC). The genetic components of nine clinical isolates were investigated using qPCR. Two genes were targeted: icaA a biofilm adhesion gene, and blaZ a common ampicillin resistance gene. icaA positive isolates, on average, were observed to form larger biofilms on catheter tips compared with icaA negative isolates, while no significant difference was observed between genotype and phenotype when assessing biofilms grown on a microtiter plate. The presence or absence of the blaZ gene played an important role in ampicillin resistance, regardless of a small or large biofilm production. A single clinical isolate, #46, which lacked the icaA gene but was observed to form a large biofilm on the microtiter plate and both commercial catheter tips. Furthermore, biofilms formed by this same isolate was observed to provide the highest BMIC resistance against ampicillin. Further analysis of the biofilm formed by isolate #46 showed that extracellular DNA (eDNA) and proteins, not polysaccharides, are the primary components of the biofilm. Finally, preliminary data is presented that describes a modified Biofilm flow-system that will assist future studies to model S. aureus biofilm formation on catheter tips that exist in a flow environment, similar to that observed in a patient.