68 / 453
4
the 3M MDA 2 –
E. coli
O157 (including H7) method were compared to samples analyzed using the USDA/FSIS
1
MLG reference method in an unpaired study design. All positive test portions were biochemically confirmed by
2
the API 20E biochemical testor by the VITEK 2 GNbiochemical identification test, AOAC Official Method
3
2011.17 [7].
4
5
Statistical Analysis
6
7
Each collaborating laboratory recorded results for the reference method and the 3M MDA 2 –
E. coli
O157
8
(including H7) method on the data sheets provided. The data sheets were submitted to the study director at the end
9
of testing for statistical analysis. Data was analyzed using the Least Cost Formulations, Ltd., AOAC Binary Data
0
Interlaboratory Study Workbook
( http://lcfltd.com/aoac/aoac-binary-v2-3.xls)provided by AOAC RI [8]. Data for
1
each test portion size was analyzed using the probability of detection (POD)statistical model [9].The probability of
2
detection (POD) was calculated as the number of positive outcomes divided by the total number of trials. The POD
3
was calculated for the candidate presumptive results, POD
CP,
the candidate confirmatory results (including false
4
negative results), POD
CC
, the difference in the candidate presumptive and confirmatory results, dLPOD
CP,
5
presumptive candidate results that confirmed positive (excluding false negative results), POD
C,
the reference
6
method, POD
R
, and the difference in the confirmed candidate and reference methods, dLPOD
C
. A
7
dLPOD
C
confidence interval not containing the point zero would indicate a statistically significant difference
8
between the 3M MDA 2 –
E. coli
O157 (including H7)and the reference methods at the 5 % probability level. In
9
addition to POD, the repeatability standard deviation (s
r
), the among laboratory repeatability standard deviation
0
(s
L
), the reproducibility standard deviation (s
R
)and the P
T
value were calculated. The s
r
provides the variance of
1
data within one laboratory, the s
L
provides the difference in standard deviation between laboratories and the s
R
2
provides the variance in data between different laboratories. The P
T
value provides information on the
3
homogeneity test of laboratory PODs [9].
4
5
AOAC Official Method
2016.xx6
Escherichia coli
O157:H7in Selected Foods
7
3M
™
Molecular Detection Assay(MDA) 2 –
E. coli
O157 (Including H7) Method
8
First Action 2016
9
0
(Applicable to detection of
Escherichia coli
O157 (including H7) in raw ground beef (73% lean), frozen
1
blueberries, fresh sprouts, and fresh baby spinach.
2
3
See Table
2016.1A
for a summary of results of the inter-laboratory study.
4
See Table
2016.2A
for detailed results of the inter-laboratory study
5
6
A.
Principle
7
8
The 3M
™
Molecular Detection Assay (MDA) 2 –
E. coli
O157 (including H7) method is used with the 3M
™
9
Molecular Detection System (MDS) for the rapid and specific detection of
E. coli
O157 (including H7) in enriched
0
foodsamples. The 3M MDA2 –
E. coli
O157 (including H7)uses loop-mediated isothermal amplification of unique
1
DNA target sequences with high specificity and sensitivity, combined with bioluminescence to detect the
2
amplification. Presumptive positive results are reported in real-time while negative results are displayed after the
3
assay is completed, 60 minutes. Samples are enriched in 3M
™
Buffered Peptone Water (BPW) ISO formulation.
4
5
B.
Apparatus and Reagents
6
Items (a)-(g) are available as the 3M
™
Molecular Detection Assay (MDA) 2 –
E. coli
O157 (including H7)
7
kit from 3M Food Safety (St. Paul, MN 55144-1000, USA).
8
OMAMAN-35 A : Collaborative Study Manuscript
For ERP Use Only
January 2017
AOAC Research Institute
Expert Review Panel Use Only