Browsing by Author "Ashley, Jon"
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Item Open Access Development of a β-Lactoglobulin sensor based on SPR for milk allergens detection(MDPI, 2018-03-27) Ashley, Jon; D’Aurelio, Roberta; Piekarska, Monika; Temblay, Jeff; Pleasants, Mike; Trinh, Linda; Rodgers, Thomas L.; Tothill, Ibtisam E.A sensitive and label-free surface plasmon resonance (SPR) based sensor was developed in this work for the detection of milk allergens. β-lactoglobulin (BLG) protein was used as the biomarker for cow milk detection. This is to be used directly in final rinse samples of cleaning in-place (CIP) systems of food manufacturers. The affinity assay was optimised and characterised before a standard curve was performed in pure buffer conditions, giving a detection limit of 0.164 µg mL−1 as a direct binding assay. The detection limit can be further enhanced through the use of a sandwich assay and amplification with nanomaterials. However, this was not required here, as the detection limit achieved exceeded the required allergen detection levels of 2 µg mL−1 for β-lactoglobulin. The binding affinities of the polyclonal antibody for BLG, expressed by the dissociation constant (KD), were equal to 2.59 × 10−9 M. The developed SPR-based sensor offers several advantages in terms of label-free detection, real-time measurements, potential on-line system and superior sensitivity when compared to ELISA-based techniques. The method is novel for this application and could be applied to wider food allergen risk management decision(s) in food manufacturing.Item Open Access Development of an immunosensor for PfHRP 2 as a biomarker for Malaria detection(MDPI, 2017-07-18) Hemben, Aver; Ashley, Jon; Tothill, Ibtisam E.Plasmodium falciparum histidine-rich protein 2 (PfHRP 2) was selected in this work as the biomarker for the detection and diagnosis of malaria. An enzyme-linked immunosorbent assay (ELISA) was first developed to evaluate the immunoreagent’s suitability for the sensor’s development. A gold-based sensor with an integrated counter and an Ag/AgCl reference electrode was first selected and characterised and then used to develop the immunosensor for PfHRP 2, which enables a low cost, easy to use, and sensitive biosensor for malaria diagnosis. The sensor was applied to immobilise the anti-PfHRP 2 monoclonal antibody as the capture receptor. A sandwich ELISA assay format was constructed using horseradish peroxidase (HRP) as the enzyme label, and the electrochemical signal was generated using a 3, 3′, 5, 5′tetramethyl-benzidine dihydrochloride (TMB)/H2O2 system. The performance of the assay and the sensor were optimised and characterised, achieving a PfHRP 2 limit of detection (LOD) of 2.14 ng·mL−1 in buffer samples and 2.95 ng∙mL−1 in 100% spiked serum samples. The assay signal was then amplified using gold nanoparticles conjugated detection antibody-enzyme and a detection limit of 36 pg∙mL−1 was achieved in buffer samples and 40 pg∙mL−1 in serum samples. This sensor format is ideal for malaria detection and on-site analysis as a point-of-care device (POC) in resource-limited settings where the implementation of malaria diagnostics is essential in control and elimination efforts.Item Open Access A rate-based approach to cleaning-in-place(American Chemical Society, 2017-05-18) Trinh, Linda; Willey, A. Roy; Martin, Peter J.; Ashley, Jon; Tothill, Ibtisam E.; Rodgers, Thomas LawrenceFouling is a major problem in the food industry. In cases where allergens are involved, excessive cleaning of food processing equipment often takes place to ensure allergen levels are reduced to acceptable levels. Experimental trials were conducted in a pilot scale cleaning rig to assess the effect of fluid flow rate and temperature on the removal rate of milk paste during cleaning-in-place. Conductivity measurements taken during trials were analysed to produce an equation enabling calculation of the cleaning rate with water temperature, flow rate, initial deposit mass, and time. The rate of cleaning with detergent was also investigated. This methodology produces a rate based expression for cleaning which can be applied to other products, and can be used to determine the time when cleaning is complete, the extent of a cleaning process, or when the pre-rinse should be replaced with a detergent rinse. This prevents over-cleaning and therefore reduces the costs associated with over-cleaning through energy, material and loss of production time.