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This study evaluated the ability of the 3M Molecular Detection Assay
Salmonella
method to remain
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unaffected by small variations in method parameters that might be expected to occur when the method is
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performed by an end user. The following effects of perturbing method parameters were investigated:
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1. Enrichment volume variation:
Following step 4.2 (Lysis) in the method instructions, 18, 20 and 22µL of
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each enriched sample were added to individual lysis tubes.
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2. Lysis time
: Following step 5 in the method instructions, the LS tubes containing the enriched sample were
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placed onto the 3M Molecular Detection Heat Block for 13, 15 and 17 min at 100°C following the protocol
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below.
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3. Lysis volume variation
: Following step 4.2 (Amplification) in the method instructions, 18, 20 and 22µL of
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sample lysate was added to individual reaction tubes.
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The volumetric variations were based on allowing an end-user to deviate (via pipettor setting, calibration
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error or erroneous technique) from the specified transfer volume by as much as 10% for enrichment &/or
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lysate. This testing determined robustness of the assay due to reasonable variations in transferred sample
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volume. NOTE: This testing measured the additive effect of extremes in both transfer volumes, e.g.,
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enrichment and lysate volumes both at -10% or both at +10%. This design was chosen based on prior
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evidence that coupling these extremes did not lead to different results, therefore it was decided this was a
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more rigorous challenge, versus testing one volumetric transfer error at a time (holding the other constant).
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Two positive isolates (
S
.
Typhimuirum
ATCC 14028 and
S
.
Enteritidis
ATCC 13076) were analyzed within one
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log of the limit of detection of the kit. Each isolate was cultured in 3M BPW ISO and then diluted in the
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same broth to within one log limit of detection of the kit. One non-
Salmonella
isolate (
Citrobacter freundii
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ATCC 8090) was analyzed at the overnight growth level achieved in a nonselective broth (BHI). Five (5)
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replicates of each isolate and a BPW control were analyzed using the 3M Molecular Detection Assay
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Salmonella
Method.
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Ruggedness Testing Results
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Results for the 3M Molecular Detection Assay
Salmonella
ruggedness study are summarized in Table 4
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in the Appendix. Both
Salmonella
strains were positive for all replicates and method variations. The
C.
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freundii
strain and BPW control was negative for all replicates and method variations. The 3M
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Molecular Detection Assay
Salmonella
method is sufficiently rugged to withstand small variations in
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the experimental parameters of enrichment sample volume, lysis time and lysis temperature.
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Shelf Life and Lot-to-Lot Consistency Methodology
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For the lot-to-lot consistency study, 3 lots of test kits (1 newly manufactured, 1 lot at the middle of its term
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and 1 lot near the expiration date) were evaluated. The target shelf-life for the 3M Molecular Detection
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Assay-
Salmonella
is 18 months. In addition to testing for lot-to-lot consistency, this phase of testing also
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analyzed the stability of the test kit, concurrently.
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Two positive isolates for
Salmonella
(
S
.
Typhimuirum
ATCC 14028 and
S
.
Enteritidis
ATCC 13076) and one
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non-
Salmonella
isolate (
C. freundii
ATCC 8090) were analyzed. Each isolate was cultured in 3M BPW ISO
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and then diluted in the same broth to within one log limit of detection of the kit.
For each isolate, eight
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replicates of were tested (16 positive and 8 negative) using the 3M Molecular Detection Assay
Salmonella
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Method.
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