Development of Microscopic Imaging System for Rapid Detection of Salmonella in Raw Chicken
Type of Degreedissertation
Nutrition and Food Science
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The overall objective of this study was to develop a new Salmonella detection method on poultry by combining a biosensor with a light microscopic imaging system. In the first study, brain heart infusion (BHI) broth was the most efficient enrichment medium for Salmonella growth which increased the Salmonella population by 4-logs in 6 h, whereas, brilliant green (BG) broth was the most efficient medium for enriching Salmonella in chilled poultry. The polyclonal antibody (pAb) was specific to entire Salmonella strains tested, except S. Diarizonae. In the second study, the gold and polystyrene sensor platforms immobilized with pAb possessed significantly high binding efficiencies with Salmonella, showing 66 ± 24 CFU/0.0013 mm2 and 57 ± 16 CFU/0.0013 mm2, respectively (p < 0.05). However, the protein A and lysine treatments on gold or polystyrene sensor platform did not enhance antibody immobilization. The optimum concentration of pAb, temperature, and the range of pH in PBS were 100 μg/mL, 30 C, and pH 7.0-8.0, respectively. The detection limit of the gold biosensor with light microscopic imaging system (GB-LMI) was 2.0 ± 1.0 CFU/0.0013 mm2 with inoculation of 103 CFU. The GB-LMI showed that the detected number of Salmonella enriched in BHI medium for 6 h with the initial population of 102 CFU and 103 CFU was significantly higher than that of enriched in BG medium, (i.e., 75 ± 15 CFU/0.013 mm2 for 6 h-103 in BHI and 51 ± 8 CFU/0.013 mm2 for 6 h-103 in BG). Therefore, the GB-LMI method could detect Salmonella on chicken skin inoculated within 4.5 h after 4 h-enrichment in BHI and BG media. In the third study, the optimum quantity of pAb for magnetic beads conjugation, time, reaction temperature, buffer type and pH were 0.44 mg per mL MBs, 20 min, 30 C, and pH 6.5-7.5 in PBS buffer, respectively. The conjunction of GB-LMI and IMS method did not increase the number of Salmonella captured on gold biosensor significantly (32.0 ± 0.9 cells/0.013 mm2 for chicken sample enriched in BHI for 6 h at the inoculation of 102 CFU/16 in2 and 74.0 ± 9.5 cells/0.013 mm2 for chicken sample enriched in BHI for 6 h at the inoculation of 103 CFU/16 in2). However, the method improved the resolution of microscopic images with enhancing the reliability and accuracy of GB-LMI detection.