Abstract
A very small number of bacterial pathogens may have fatal effects on food safety. In spite of having great advancements in bioanalytical methods, most of the accepted detection methods are still cultivation based and thus time consuming. This leads to an intense need for efficient and rapid methods for detection of food-related bacteria. In this study, a flow cytometry based immunomagnetic separation (IMS) method for the isolation and enrichment of Salmonella Typhimurium from liquid samples was developed and optimized. Both polyclonal and monoclonal antibodies have been used to couple with 1 micron sized paramagnetic particles for the preparation of immunomagnetic beads (IMBs). The most suitable antibody was chosen by applying an enzyme linked immunosorbent assay (ELISA), whereas living bacteria were detected by flow cytometry. The parameters for both IMS and flow cytometry e.g., concentration of bead and bacteria, immunocapture time, staining and buffering conditions for the viability assays were optimized. The capture efficiency of IMS was>98% for a range of Salmonella Typhimurium cell concentrations from 103 to 105/mL using 108/mL bead concentration. The method proved to have high (98%) specificity towards Salmonella Typhimurium and very low (< 5%) binding with non-target bacterial strains.
Highlights
Due to an increased number of recalls and cases involving microbial contamination and adulteration of food, food processing industries require on-line fast, sensitive and early detection methods for microbial contaminants to avoid withdrawing a large amount of products from the market
The most suitable antibody was chosen by applying an enzyme linked immunosorbent assay (ELISA), whereas living bacteria were detected by flow cytometry
The efficiency of immunomagnetic separation (IMS) mainly depends on the affinity of the antibody towards the target bacterium that is used for the preparation of immunomagnetic beads (IMBs)
Summary
Due to an increased number of recalls and cases involving microbial contamination and adulteration of food, food processing industries require on-line fast, sensitive and early detection methods for microbial contaminants to avoid withdrawing a large amount of products from the market. Both pathogenic bacteria and non-pathogenic spoilage microorganisms cause severe problems in the food industry. Used analytical procedures in food industries are dependent on traditional culture based methods that include enrichment, plating on selective differential agar and biochemical confirmation (USDA, 2004; USFDA, 2002). The necessity for capturing low numbers of specific microorganisms in food sample results in the need for efficient techniques like immunomagnetic separation (IMS), which offers several advantages over traditional culture enrichment processes
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