AOAC ERP MICRO AUGUST 2018

17.9.24

will bind to Salmonella antigens if they are attached to adsorbed antibody on surface of well. Wells are washed to remove unbound conjugate, and enzyme substrate is added. Dark blue-green color indicates presence of Salmonella antigen. B. Reagents Items ( a )–( k ) are available as 3M TECRA Salmonella Visual Immunoassay (VIA). Items ( a )–( l ) are available as 3M TECRA Salmonella ULTIMA Immunoassay. Item ( m ) is available as TMBMED500 from 3M Microbiology, 13 Rodborough Rd, Frenchs Forest, NSW2086, Australia; www.tecra.net; and 3MMicrobiology, 3M Center, Bldg 275-5W-05, St. Paul, MN 55144-1000, USA; www.3M.com/microbiology. Substitutions must be pretested for equivalency. ( a ) Antibody adsorbed strips .—Microplate well strips, coated with polyclonal antibodies to Salmonella 96 wells. Store wells at 2 ° –8 ° C when not in use. ( b ) Tray .—Sufficient to secure individual wells or strips. ( c ) Control antigen .—Positive control (lyophilized). Purified Salmonella antigen, which reacts with antibodies to Salmonella , 1 vial. Reconstituted control antigen is stable 2 months when stored at 2 ° –8 ° C. ( d ) Controls diluent .—One vial (6 mL/vial). Contains 0.006 g tris [tris(hydroxymethyl)aminomethane], 0.044 g NaCl, 0.0025 g Tween 20 (polyoxyethylene 20 sorbitan monolaurate), and 0.005 g thimerosal in H 2 O. Controls diluent also serves as negative control. ( e ) Conjugate .—Two vials (lyophilized). Contains ca 150 ng anti- Salmonella antibodies (from sheep) conjugated to horseradish peroxidase, 0.00686 g Na 2 B 4 O 7 × 10H 2 O, 0.12 g Sigma

AOAC Official Method 998.09 Salmonella in Foods

Colorimetric Polyclonal Enzyme Immunoassay Screening Method with Rappaport-Vassiliadis (R10) Broth and/or Tetrathionate Broth 3M ä TECRA ä Salmonella Visual Immunoassay (VIA) First Action 1998 Revised First Action 1999 Final Action 2001 3M ä TECRA ä Salmonella ULTIMA ä Immunoassay Revised First Action 2003 Final Action 2009

(Method is screening procedure for presence of Salmonella in all foods; it is not a confirmatory test because polyclonal antibodies used in test may cross-react with small percentage of non- Salmonella organisms. Tests positive by the immunoassay must be confirmed by biological and serological procedures.) See Tables 998.09A–D for the results of the interlaboratory studies supporting acceptance of the method. A. Principle Detection of Salmonella antigens is based on enzyme immunoassay using highly purified antibodies prepared from antigens unique to Salmonella . Polyclonal antibodies to Salmonella antigens are adsorbed onto internal surface of 96-well microtiter tray. Broth to be tested is placed into a well of a tray. If Salmonella antigens are present, they will attach to specific antibody adsorbed on well. All other material is washed away. Conjugate is added and

Table 998.09A. Statistical analyses of interlaboratory study results for 3M TECRA assay using 2 selective enrichment broths

Incidence of false negatives among

Incidence of false positives among total negative samples, % f VIA Culture

total positive samples, % d

Sensitivity rate, % e

Specificity rate, % g VIA Culture 99 91 100 98 100 98 95 100 99 100 100 100 100 100 100 100 100 96 100 99 100 99 100 99 99 99 100 100 95 45 96.8 92.1 100 95.1

MPN a (CFU/g)

Method agreement, % b

c 2 c — h

Food (No. labs)

VIA Culture

VIA Culture

Milk chocolate (14) <0.003

100 100 100 100

— — — — 1.4

8.6 2.3 2.1 0.0 0.0 0.0 0.0 0.0 4.0 0.9 1.2 1.3 1.2 0.0

0.009 0.093

— 0.0

0.0 0.0

100 100 100 100

0.0 0.0

0.0

0.0

Dried whole egg (13) <0.003

— — — — — 4.6

0.009 98.4

0.0

0.0

2.8 0.0

100 97 100 100

1.1 0.0

0.430

100 100

— 0.0

Nonfat dry milk (18) <0.003

— — — — — 0.0

0.24

98.8

0.0

1.3

0.0 0.0

99 100 0.0 (1.2r) 0.0 100 (99r) 100

2.4

100 100 100 100 100

— 0.0

100 100

0.0

Black pepper (15) <0.003

— — — — — 0.0

0.014 0.088 <0.003

— 0.0 — 0.0

0.0

100 100

0.0 0.0

0.0 100 (97r) i 100

Soy flour (15)

— — — — — 0.0

0.24

98.7

— — 0.0 — — 0.0

100 100 100 100

1.2 0.0

2.4

100 100

Raw turkey (12) (BAM, 7th Ed.)

<0.003

— — — — — 5.0 55

0.043 93.3

0.0 1.3

1.8 0.0

5.3 5.1

98.2 94.7 100 94.9

3.2 0.0

7.9 4.9

0.46

95.0

a–i See Table 998.09D for footnotes.

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