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detected using a specific molecular beacon probe. Fluorescent signal is generated in real time, with
amplification and detection complete within 10 minutes. The entire assay is conducted at a constant
temperature of 56
o
C using a temperature-controlled fluorescence detection instrument. Assay software
analyzes the fluorescent signal over time; a data interpretation algorithm interprets results as negative,
positive, or invalid based on baseline, rate-of-change, and other criteria. Each tube of ANSR reagents
also contains an internal positive control, signaling in a second fluorescence channel irrespective of the
presence of target DNA, and indicating proper functioning of the amplification reagents.
B.
Media and Reagents
(a)
ANSR
TM
for Salmonella test kit. –
Available from Neogen Corp. (Lansing, MI,
www.neogen.com ).Contains: – Lyophilized reagents in capped strip tubes, 8 tubes per strip, 12 strips
(96 tests) per kit, in 2 sealed foil pouches with desiccant packs;cluster tubes, 8 tubes per strip, 12 strips
per kit; permanent caps, 8 caps per strip, 12 strips per kit; lysis buffer, 1 bottle, 50 mL; Kit insert.Store
reagent tubes at 2-8
o
C, in sealed foil pouches with desiccant. Store lysis buffer at 2-8
o
C.
(b)
Phosphate-buffered saline
. – Per liter: 8.0 g NaCl, 0.2 g KCl, 1.44 g Na
2
HPO
4
, 0.24 g KH
2
PO
4
.
(c)
Hektoen enteric agar
. – Available from Neogen Corp. and other suppliers. Follow
manufacturer’s instructions for preparation.
(d)
Xylose lysine deoxycholate (XLD) agar
. – Available from Neogen Corp. and other suppliers.
Follow manufacturer’s instructions for preparation.
(e)
Bismuth sulfite agar
. – Available from Neogen Corp. and other suppliers. Follow manufacturer’s
instructions for preparation.
(f)
Brilliant green sulfa agar
. – Available from Neogen Corp. and other suppliers. Follow
manufacturer’s instructions for preparation.
(g)
Xylose lysine tergitol (XLT4) agar
. – Available from Neogen Corp. and other suppliers. Follow
manufacturer’s instructions for preparation.
(h)
Double-modified lysine iron agar
. – Available from various suppliers. Follow manufacturer’s
instructions for preparation.
(i)
Tryptic soy agar
. – Available from Neogen Corp. and other suppliers. Follow manufacturer’s
instructions for preparation.
C.
Apparatus
3