| dc.description.abstract | Staphylococcus aureus contamination in milk demands swift detection methods.
Lateral Flow Immuno Assays (LFIA) show promise but struggle with milk's complexity.
To overcome this, an LFIA model using Anti-Staphylococcal �� -Toxin antibody for
S.aureus detection is proposed. This study focuses on developing sample processing
methods tailored for molecular assays, addressing challenges posed by whole milk.
Solutions involve strategic pretreatment and dilution techniques to optimize LFIA efficacy,
emphasizing specificity, sensitivity, and practicality for laboratory applications.
Staphylococcus aureus cultivation followed standardized protocols, with subsequent
molecular confirmation via Polymerase Chain Reaction (PCR) and verification of alpha
toxin production through hemolysis activity on blood agar. These methodologies
confirmed the presence of the hla gene encoding �� -Toxin in SA and demonstrated its
ability to produce this toxin, as evidenced by observed hemolysis activity on blood agar
plates. Innovative sample processing methods, such as enhanced filtration systems and
surfactant additions, are being investigated to refine the performance of molecular assays
in whole milk. Preliminary results from employing an enhanced filtration system for
pathogen detection bacterial samples are encouraging. The theoretical recovery rate of
78.7% closely mirrors the actual recovery rate of 74.6%, demonstrating the effectiveness
and efficiency of the system. Incorporating Tween, a surfactant, into the sample processing
workflow significantly enhances pathogen detection efficiency. When introduced into PBS
to dilute the milk sample, Tween has demonstrated superior performance compared to
undiluted milk samples or dilution with conventional PBS alone. Dilution with Running
Buffer and Tween (0.1% and 0.05%) has markedly improved flow times compared to nonpretreated milk samples (p-value < 0.05). Statistical analyses underscore the pronounced
impact of Tween concentration on LFIA performance. Comparative assessments reveal the
efficacy of PBS + 0.1% Tween and PBS + 0.05% Tween at 1:1 dilution in mitigating milk
matrix effects and enhancing flow dynamics with no significant differences between these
two concentrations of Tween. The investigation into the Direct Addition of Tween to milk
samples presents compelling evidence of its superior performance in optimizing the LFIA diagnostic process, especially when utilizing 0.1% Tween concentration. The exploration
of the LFIA full-strip model, incorporating novel targets and antibodies, offers valuable
insights into the efficacy of various milk pretreatment methods, including dilution with
PBS containing Tween and direct addition of Tween. Significantly, the dilution with PBS
containing 0.1% Tween emerges as the most promising approach, yielding valid results
with clear C and T lines, in contrast to other methods hampered by milk-induced flow
obstructions. These findings underscore the optimal method for detecting SA in milk
matrices, emphasizing the importance of dilution with PBS containing 0.1% Tween.
Moreover, this study contributes valuable insights to the field, guiding future advancements
in pathogen detection methodologies for dairy products. | en_US |
