Singlepath® Salmonella

EMD Cat. No. 1.04140.0002
25 tests


GLISA-Rapid Test (Gold Labelled ImmunoSorbent Assay) for the qualitative detection of Salmonella spp. in food.

Introduction
 Salmonella is one of the most common causes of food poisoning world-wide. Salmonella have been isolated from most types of raw food (meats, eggs, as well as plant products), and their high resistance to drying combined with a very high heat resistance once dried, makes Salmonella a potential problem in most foods and in particular in dried products.
 Screening for the presence of Salmonella in foods by conventional methods involve a 3-step technique: non-selective pre-enrichment (18-24 h), selective enrichment in (at least) two different selective broth media (24-48 h) followed by plating on (at least) two different selective/indicative agars (24-48 h).This leads to a total time for Yes/No screening result of up to 5 days. For products where a positive release system is enforced, this means a delay of 5 days before release of the product.
 The Singlepath® Salmonella test is performed only from one selective enrichment culture, and gives a Yes/No answer in 20 minutes, meaning that products can be released 2 days earlier than by conventional microbiology.
Principle
 Singlepath® Salmonella is an immunochromatographic rapid test based on gold-labelled antibodies. The test device has a circular sample port, and an oval shaped test (T) and control (C) window.
 1. The sample is applied to the chromatography paper via the circular sample port
 2. The sample is absorbed through the pad to the reaction zone containing colloidal, gold-labelled antibodies specific to Salmonella spp.
 3. Any Salmonella antigen present complexes with the gold-labelled antibody and migrates through the port until it encounters a binding zone in the test (T) area.
 4. The binding zone (T) contains another anti-Salmonella Antibody, which immobilises any Salmonella-antibody complex present. Due to the gold-labelling, a distinct red line is then formed.
 5. The rest of the sample continues to migrate to a second binding reagent zone within the control (C) zone, and also forms a second distinct red line (positive control). Regardless of whether any Salmonella is present or not, this distinct red line is always formed in the control (C) zone, thus ensuring the test is working correctly.
Storage/Stability
 Singlepath® Salmonella is stable until the expiry date printed on the box, when stored at +2 to +8 °C.
Materials required for the test
 Package contents
 25 test devices (individually pouched in aluminium foil)
 Additionally required materials and instrumentation
 1. Enrichment media, e.g. 1.07228. Buffered Peptone Water, 1.07700. Salmonella Enrichment Broth acc. to Rappaport-Vassiliadis (RVS broth).
2. Stomacher / Stomacher bags
 3. Incubators +37°C and +41.5°C
 4. Distilled or deionized water
 5. Autoclave
 6. Waterbath for boiling of samples
 7. Disposable heat-stable Polypropylene tubes for boiling of samples
 8. Disposable plastic transfer pipettes and/or appropriate micro pipettes and disposable tips for dispensing 1-2 ml (sample for boiling) and 160 µl (application of boiled sample onto tests)
Sample material / sample enrichment
 Mix 25 g solid sample or 25 ml liquid sample with 225 ml pre-enrichment broth (BPW) and homogenise in stomacher if necessary.
 Incubate for 18-24 h at 37 °C.
 Inoculate 9.9 ml RVS selective enrichment broth with 0.1 ml of pre-enrichment culture. Incubate for 24 h at 41.5 °C. (If a different selective enrichment broth is used, follow recommendations for incubation time/temperature).
Test procedure
 Sample Preparation
 1. Transfer approx. 1-2 ml of selective enrichment culture to an appropriate (polypropylene) tube.
 2. Place tubes in boiling water bath for 15 min.
3. Remove and allow cooling to room temperature.
 Procedure
 1. Remove the foil pouches from the required number of Singlepath® Salmonella devices. Place the test device(s) on a flat surface and label with appropriate sample identification. (Note: Perform the tests within a period of 2 hours after opening!)
 2. Using a disposable transfer pipette, draw up a sample from the boiled, cooled enrichment. Squeeze the pipette bulb, insert the stem into the boiled sample and release pressure on bulb. This will draw sample up into the pipette. Dispense five (5) free falling drops (about 150-160 µl) into the circular sample port on the test device.
Alternatively using a micro pipette and disposable pipette tip, dispense 160 µl sample into the circular sample port on the test device.
 3. Observe the test result 20 minutes after applying the sample to the device.
 Interpretation of results
 The test can be regarded as working correctly if a distinct red line appears in the control zone (C) within 20 minutes.
 A sample can be considered POSITIVE if at or prior to 20 minutes, red lines appear on both test (T) and control (C) zones.
 A sample can be considered NEGATIVE if no red line appears in the test (T) zone but does appear distinctly in the control (C) zone 20 minutes after application of sample to the device.
Technical specifications
 Detection limit
 Depending on serogroup, a range of approx. 104 107 bacteria/ml can be regarded as being the lower detection limit.
 Interferences
 Results obtained to date on numerous food samples indicate that there is no interference of Singlepath® Salmonella with food ingredients.
 The test has been developed based on using RVS and TTB selective enrichments and Merck M-broth.
Interference from other types of selective enrichment broth and other brands of M-broth cannot be excluded. In particular use of broth of red-brown color could potentially mask weak signals due to background coloration of the test zone.
Trouble-shooting
Problem
 		Measures
No line appears in either zone after 20 minutes test period.
Re-run sample.
Delay in sample reaching Nitrocellulose membrane
Touch sample pad with pipette tip







Picture/Image



Singlepath® Salmonella



© 2002 Merck KGaA, Darmstadt, Germany