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All of the assay steps are performed automatically by the instrument. The reaction
1
medium is cycled in and out of the SPR® several times. Part of the enrichment broth is
2
dispensed into the reagent strip. The receptors present will bind to the proteins specific
3
for
Listeria
receptors, which are coated on the interior of the SPR®. Unbound sample
4
components are washed away. Protein conjugated with alkaline phosphatase are cycled in
5
and out of the SPR® and will bind to any
Listeria
receptors which are in turn bound to
6
the specific protein on the SPR® wall.
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Further wash steps remove unbound conjugate. During the final detection step, the
9
substrate (4-Methylumbelliferyl phosphate) is cycled in and out of the SPR®. The
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conjugate enzyme catalyzes the hydrolysis of this substrate into a fluorescent product (4-
11
Methylumbelliferone), the fluorescence of which is measured at 450 nm.
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At the end of the assay, the results are analyzed automatically by the instrument which
14
generates a test value for each sample. This value is compared to a set of stored standards
15
(thresholds) and each result is interpreted (positive, negative).
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General Information
18
The current classification of the genus
Listeria
includes six species that are non-spore-
19
forming, short, motile, Gram positive rods. These organisms are catalase positive, are
20
oxidase negative, hydrolyze esculin and ferment glucose without gas production. They
21
can grow at wide temperature and pH ranges and can tolerate high concentrations of
22
sodium chloride (1,2).
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Members of the genus
Listeria
are ubiquitous and certain species are pathogenic.
Listeria
25
have been isolated from various food products including dairy products, meats,
26
vegetables and seafood, as well as from environmental samples taken, in particular, from
27
food processing plants.
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In humans, Listeriosis is usually associated with the species
L. Monocytogenes
and may
30
cause meningitis, septicemia, encephalitis, and abortions. Groups at greatest risk include
31
pregnant women, neonates, immunocompromised patients and the elderly (3).
32
Traditional methods for the detection of
Listeria
involve enrichment that includes several
33
tedious and labor intensive steps. In addition, current culture methods require lengthy
34
incubation periods to achieve final results (4,5).
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36
Methods and Materials
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