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AOAC INTERNATIONAL
A second set of samples for culture method testing using the Health Canada Compendium method
395
was collected by sponge hydrated with 10mL DE broth. After returning the sponge to the sample
396
bag, samples were held at 4±3°C for 2-3 hours prior to addition of enrichment media. Sponges
397
were mixed (by stomaching for 2 min at 200 rpm in a sterile Stomacher bag) in 225 mL of BPW
398
broth. Using 1 N HCl or NaOH, pH was adjusted if necessary to 6.0-7.0. Samples were incubated
399
for 18-24 hours at 35
o
C and subjected to BAX® System assay before secondary enrichment in
400
RVS and TVG broths. Additionally, for each sample, a 3 hr regrowth in BHI (10μl of enriched
401
sample added to 500μl of prewarmed BHI which was incubated at 37°C for 3 hrs) was performed
402
and BAX® testing was assayed on this sample as well in order to have a validated protocol for
403
samples where PCR inhibitors may be present on the sampled surfaces. All samples were then
404
struck in 10-μL amounts on BS and BGS agars. Agars were incubated at 35°C for 24±2 hr after
405
which plates were examined for colonies with typical characteristics of
Salmonella
. Suspect
406
colonies were confirmed using the biochemical and serological methods described in the Health
407
Canada Compendium.
408
409
BAX® System Assay
410
Follow prompts in the Rack Wizard to enter identifying data on the entire rack and on the individual
411
samples.
412
Place one cluster tube per sample in a cluster tube rack. Add 200
µ
L of prepared lysis reagent to
413
each tube (lysis tube). Transfer 5
µ
L of each pre-enriched sample to the corresponding lysis tube
414
and ensure the tubes are capped. Incubate lysis tubes at 37ºC for 20 minutes, then at 95ºC for 10
415
minutes in either individual heating blocks or the DuPont Thermal Block. Cool lysates for at least
416
five minutes in either a refrigerated cooling block or the automated thermal block prior to the final
417
transfer to PCR tubes.
418
Warm up the cycler/detector by selecting RUN FULL PROCESS from the menu bar of the
419
application window.
420
Dupont BAX Salmonella PTM Report
PTM Certification No. 081201 (08/07/2012)
For Expert Review Panel Use Only
Do Not Distribute