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VIDAS LMX & LPT Collaborative Study
April 19, 2013
OMA-2013-Apr-XXX
2.1
25g:
Add 225 mL LPT broth (prewarmed at 18-25°C) to each 25g test portion.
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Stomach each sample for approximately two minutes and incubate 26-30 h at 30 ±
2
1°C.
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2.1.1
Heat and Go: Transfer 0.5 mL to VIDAS LPT strip and heat on the Heat
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and Go for 5±1 min. Cool 10 min then perform VIDAS assay.
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2.1.2
Boiling: Transfer 1-2 mL of enrichment broth into a tube. Seal tube and
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heat for 5±1 min at 95-100°C. Cool, mix and transfer 0.5 mL into the
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VIDAS LPT strip then run the VIDAS assay.
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2.2
125g:
Add 375 mL LPT broth (prewarmed at 18-25°C) to each 125g test portion.
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Stomach each sample for approximately two minutes and incubate 24-30 h at 30 ±
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1°C. Transfer 1 mL primary enrichment into 10 mL LPT broth. Incubate 22-26
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hr at 30 ± 1°C.
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2.2.1
Heat and Go: Transfer 0.5 mL to VIDAS LPT strip and heat on the Heat
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and Go for 5±1 min. Cool 10 min then perform VIDAS assay.
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2.2.2
Boiling: Transfer 1-2 mL of enrichment broth into a tube. Seal tube and
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heat for 5±1 min at 95-100°C. Cool, mix and transfer 0.5 mL into the
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VIDAS LPT strip then run the VIDAS assay.
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2.3
Confirmation
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2.3.1
Confirm each test portion, regardless of presumptive result.
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2.3.2
Streak each enriched test portion to OX and ALOA following incubation
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instructions.
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2.3.3
Choose 5 typical or suspect colonies, or if there are <5 such colonies,
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choose all to confirm
Listeria
. Streak chosen colonies individually onto
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surfaces of separate tryptic soy agar with yeast extract agar plates.
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Incubate 24 hr at 37
°
C.
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2.3.3.1 Only one colony needs to confirm positive to call the test portion
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positive for
Listeria.
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2.3.4
Biochemical tests will be performed using the VITEK2 GP or API
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Listeria. Follow instructions described in AOAC 993.12 (Appendix 9.7)
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for additional tests.
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2.3.4.1 VITEK2 GP: perfom Gram stain (Gram +) followed by VITEK2
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GP card.
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2.3.4.2 API Listeria: perform Gram stain (Gram +), catalase (+), oxidase (-
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), hemolysis and motility (motile at 25
°
C) tests followed by the
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API Listeria strip.
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3.0
AOAC 993.12
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3.1
Add each 25g sample into 225 mL selective enrichment medium prewarmed to
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45°C. Blend each sample for 1-3 min according to AOAC 993.12 and incubate for
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48 h at 30
°
C
.
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3.2
If necessary, adjust
pH to 7.3 at 25 ± 0.1°C, using 1M HCl or NaOH solutions.
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3.3
Streak a loopful (~10µL) of each enriched sample onto Oxford medium,
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restreaking with fresh loops 2-3 times so well-isolated colonies can develop.
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Invert and incubate plates 48 h at 37
°
C.
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DRAFT DOCUMENT
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