130 / 1195
C.
Media
1
a)
BAX® System MP media – DuPont catalog D12404925 (bulk powder) or D12745725 (StatMedia)
2
b)
Brain Heart Infusion (BHI) broth – Oxoid catalog CM 1032 or equivalent
3
c)
Brilliant Green Water – Dissolve 1 g brilliant green dye in 10 mL sterile distilled water, then
4
dilute to 100 mL. Add 2 mL of this brilliant green dye solution to 1,000 mL sterile distilled water.
5
d)
Buffered Peptone Water (BPW) – Oxoid catalog CM 0509 or equivalent
6
e)
Lactose Broth (LB) – Dissolve 3 g of beef extract, 5 g of peptone and 5 g of lactose into 1 L of
7
distilled water. Dispense into containers and autoclave 15 minutes at 121°C. Final pH is 6.9±0.2.
8
f)
Tryptic Soy Broth (TSB) - DIFCO catalog 211825 or equivalent
9
g)
Modified Tryptic Soy Broth with novobiocin (TSB+n) - DIFCO catalog 211825 or equivalent, plus 2
10
mg/L novobiocin
11
h)
Modified Tryptic Soy Broth with casamino acids and novobiocin (TSB+caa+n) - DIFCO catalog
12
211825 or equivalent, plus 10 g/L casamino acids and 8 mg/L novobiocin
13
i)
Reconstituted Nonfat Dry Milk - Suspend 100 g dehydrated nonfat dry milk in 1 L distilled water
14
and swirl until dissolved. Autoclave at 121°C for 15 minutes.
15
j)
Universal pre-enrichment broth –DIFCO catalog 223510 or equivalent
16
k)
HCl or NaOH – for adjusting pH, if necessary
17
l)
1% aqueous brilliant green dye solution – Dissolve 1 g brilliant green dye in 10 mL sterile distilled
18
water, then dilute to 100 mL. Before use, test for toxicity with positive and negative test
19
microorganisms.
20
D.
Sample Enrichment
21
Ground beef, ground beef with soy, beef trim (25 g)
– Weigh 25 g test portion into sterile container. Use
22
a stomacher, B(n), to homogenize sample for 2 minutes with 225 mL pre-warmed (35°C) BPW, C(d).
23
Incubate, B(m), at 35°C for 20-24 hours.
24
Ground beef (375 g)
- Weigh 375 g test portion into sterile container. Use a stomacher, B(n), to
25
homogenize sample for 2 minutes with 1500 mL pre-warmed (35°C) mTSB+n, C(g). Incubate, B(m), at
26
35°C for 22-26 hours.
27
Ground beef with soy (325 g)
- Weigh 325 g test portion into sterile container. Use a stomacher, B(n), to
28
homogenize sample for 2 minutes with 975 mL pre-warmed (35°C) mTSB+caa+n, C(h). Incubate, B(m), at
29
35°C for 20-24 hours.
30
Beef trim (325 g)
- Weigh 325 g test portion into sterile container. Hand massage to homogenize sample
31
for 2 minutes with 1500 mL pre-warmed (41°C) BAX® System MP media, C(a). Incubate, B(m), at 39-42°C
32
for 16-24 hours.
33
Frankfurters (325 g)
- Weigh 325 g test portion into sterile container. Use a stomacher, B(n), to
34
homogenize sample for 2 minutes with 1400 mL pre-warmed (35°C) BPW, C(d). Add additional BPW to
35
reach a total media volume of 2925 mL. Incubate, B(m), at 35°C for 18-24 hours.
36
Shrimp, peanut butter (25 g)
– Weigh 25 g test portion into sterile container. Use a stomacher, B(n), to
37
homogenize sample for 2 minutes with 225 mL pre-warmed (35°C) LB, C(e). Let stand at room
38
Dupont BAX Salmonella Pre- Collaborative Study
For Expert Review Panel Use Only
Do Not Distribute
AOAC INTERNATIONAL