Skip to 0 minutes and 1 second Lab experiment.
Skip to 0 minutes and 5 seconds A spiking experiment is set up in the laboratory as follows. 25 grammes of a pork sample is weighed and homogenised to with 225 mils diluting buffer in a Stomacher instrument.
Skip to 0 minutes and 28 seconds 10 of the 20 tubes are selected at random, and spiked with salmonella typhimurium culture.
Skip to 0 minutes and 37 seconds The other 10 samples are not spiked.
Skip to 0 minutes and 44 seconds Well 1 contains magnetic beads plus 1 mil of homogenised spiked or non-spiked pork sample.
Skip to 0 minutes and 53 seconds 1 mil of each of the 20 samples is then processed by Immunomagnetic Separation, or IMS, using the bead retriever instrument.
Skip to 1 minute and 3 seconds Mixing takes place for 30 minutes, during which time any salmonella present bind to the peptide-coated magnetic beads. Then the magnet is switched on, and beads are captured from the sample for transfer to well two.
Skip to 1 minute and 20 seconds Well 2. Beads are released into one mil of wash buffer, and washed for one minute with mixing, to remove food components and other non-target bacteria from the magnetic beads.
Skip to 1 minute and 35 seconds The magnet is then switched on, and beads are captured from the sample for transfer to well three.
Skip to 1 minute and 43 seconds Well 3 is a second wash. Same process as in well 2.
Skip to 1 minute and 51 seconds The magnet is switched on. And beads are captured from the sample for transfer to well four.
Skip to 1 minute and 58 seconds Well 4. Beads are released into the final resuspension buffer, which for PCR is Tris-EDTA buffer.
Skip to 2 minutes and 10 seconds At the end of IMS, magnetic beads are resuspended in 100 microliters of buffer.
Skip to 2 minutes and 30 seconds DNA is extracted from any captured salmonella cells by heating samples at 100 degrees C for 10 minutes in the heating block.
Skip to 2 minutes and 45 seconds Salmonella-specific PCR is carried out on all samples. And the presence or absence of the expected PCR product is recorded after agarose gel electrophoresis.
Lab experiment: IMS-PCR testing for salmonella in pork
Immunomagnetic separation (IMS) is a technique used in food testing laboratories to isolate foodborne pathogens such as Salmonella spp. from food samples.
It permits the capture and concentration of a target pathogen by means of microscopic magnetic beads coated in either target-specific antibody or peptide binders.
The video demonstrates the application of IMS to facilitate the extraction of Salmonella cells within meat samples and subsequent quantification of the level of contamination by polymerase chain reaction (PCR) analysis.
If you have any comments about the method, or if you have used this technique before please share your thoughts in the discussion area.