| Popis: |
Horseracing is a huge industry both nationally and internationally. With large financial rewards now available at some meetings the pressure to excel has grown and as a result the desire to win has led some racehorse owners to resort to the use of performance enhancing drugs. Rumours concerning the abuse of recombinant human erythropoietin (rHuEPO) in thoroughbred horseracing have been circulating since it became readily available in the late 1980s. While there are a number of commercially available EPO enzyme-linked immunosorbent assays (ELlSAs) to detect the administration of rHuEPO to horses, these kits are all restricted in their success due to the limited amount of time EPO remains in the horse's circulation. It is thought that the horse's immune system recognises the rHuEPO as a foreign protein and produces antibodies against it. This research aims to exploit this phenomenon to develop a new sensing system for the detection of rHuEPO antibodies resulting from rHuEPO administration. This research was divided into two parts. In the first section, a simple colorimetric assay based on the aggregation of gold nanoparticles for the detection of the antibodies produced as a result of rHuEPO abuse was developed. Various forms of rHuEPO have been attached to gold nanoparticles via linkage molecules to develop colorimetric assays based on the aggregation of protein-modified gold nanoparticles and its corresponding antibody. The 17 nm protein-stabilised nanoparticles are ruby red in colour. The sensing system was tested with the anti-HuEPO antibody. Upon addition of this antibody, aggregation occurred, with a subsequent change in colour from red to purple due to a red-shift in the surface plasmon absorption band, which was monitored by UV/visible spectrophotometry. For each variant of rHuEPO, the theoretical limit of detection (LOD) was established and kinetic studies (time to complete the aggregation reaction) were investigated. |
