Development and Evaluation of Colloidal Gold Lateral Flow Immunoassays for Detection of Escherichia Coli O157 and Salmonella Typhi

Lateral flow immunoassays (LFIAs) are rapid and simple detection methods for detecting bacterial contamination in food and drinking water such as Escherichia coli O157 and Salmonella Typhi. Escherichia coli O157 can cause haemorrhagic colitis, diarrhoea and haemolytic uraemic syndrome, whereas Salmonella Typhi is responsible for typhoid fever in human. Colloidal gold nanoparticles (AuNPs) synthesized using citrate reduction method is commonly used as labels for LFIAs. However, in this study, AuNPs synthesized using the seeding-growth method was evaluated as label in LFIAs. The 15 nm AuNPs seed was first synthesized using the citrate reduction method followed by growth to 40 nm AuNPs using mild reduction agent, hydroxylamine. This monodispersional AuNPs was then conjugated with anti-gram-negative endotoxin antibody at 3 μL/mL. The conjugate then immersed to conjugate pad of glass fiber at OD 10. Two sets of nitrocellulose membrane (NC) were used to immobilize anti-E. coli O157 antibody as the test dot at 1.7 mg/mL and anti-mouse IgG antibody as the control dot at 1.0 mg/mL. Another set of NC immobilized with 1.0 mg/mL anti-Salmonella sp. antibody as the test dot and the same control dot as first NC set. Spiked water and milk sample was prepared by adding 10-fold dilution ATCC stains of E. coli O157 and Salmonella Typhi to water and milk samples. LFIA to E. coli O157 in water and milk sample was able to detect as low as 7.8 x 10⁵ and 3 x 10⁶ CFU/mL, respectively, whereas LFIA to Salmonella Typhi was able to detect as low as 3 x 10⁸ and 3 x 10⁷ CFU/mL in water and milk sample, respectively. Specificity for both strips sets was 100%. This method is very useful for monitoring bacterial contamination in food and drinking water towards ensuring food and water safety. (paper)