Next Generation of Ultra-Sensitive DNA Diagnostics for Gastric Infections

We have been working on methodologies to detect DNA sequences relating to infectious disease for a number of years with a focus on using functionalised nanoparticles and surface enhanced Raman scattering. This work has produced a number of different assay approaches which are now under development through Renishaw Diagnostics Ltd. Now that these assays are moving into the hands of the end user, the market feedback suggests that improvements can be made to the assays to make them more competitive with the latest developments in the sector and also to improve the ease of use for non-specialists. This PhD studentship is designed to investigate the detection of specific DNA sequences indicative of gastric infections in a multiplexed and simple manner without the need for PCR amplification of the targets as is used in a majority of commercially available assays. The challenge of moving to a PCR free based assay is mainly in terms of the sensitivity requirements where PCR greatly increases the chance of detection but also in terms of the simplicity of hybridisation of the target with the probes through the control of the length of the DNA using PCR The aim of this proposal is to develop a new assay to detect DNA sequences in a clinically relevant range (less than 107 copies) for gastric infections without any enzyme steps. The objectives are:

1. To investigate the use of a stationary phase such as nitrocellulose for the immobilisation of a probe sequence of DNA to capture the target DNA and wash away excess. This will also involve the addition of DNA functionalised nanoparticles which we will use to hybridise to the captured target sequence on the surface. We will take a further leap in sensitivity by using a nanoparticle assembly approach which we have already shown to work through DNA hybridisation. This will take place only on the nanoparticles attached to the DNA sequence on the surface.

2. Having tested the above system we will move towards clinical samples and evaluate the performance of this assay format with clinical samples which contain a multitude of components not found in the model test systems.

3. To create a working device for verification and validation of this approach for use in ultrasensitive DNA detection which should be possible in less than 1 hour from sample collection.