NATIONAL FOOD SAFETY FOUNDATION

Transcription

1 NATIONAL FOOD SAFETY FOUNDATION RAPID TESTING KITS DETECTION E.COLI 0157 IN FOOD MATICES AND LIQUIDS USING E.COLI RAPID TESTING KIT

2 INTRODUCTION E. coli serotype O157 is a gram-negative rod-shaped bacterium. While most strains are harmless and normally found in the intestines of mammals, the O157 serogroup is a member of a class of pathogenic E. coli known as enterohemorrhagic E. coli or EHEC. Members of this class may produce Shiga-like toxins (Verotoxin), which causes severe illness particularly in those with immune deficiency, young children and the elderly. People infected with enterohemorrhagic E. coli often develop hemorrhagic colitis and/or hemolytic-uremic syndrome. The main sources of contamination include raw ground beef, unpasteurized milk, untreated water, juice, raw sprouts, lettuce, and salami. The reservoir for enterohaemorrhagic E. coli is feces of cattle, sheep and goats. The detection of E. coli O157 in food samples is often laborious and time-consuming. Several days are required to culture the bacteria from a sample followed by a final isolation and identification by biochemical and serological techniques. The E. coli O157 Strip Test Kit provides a simple, rapid, sensitive and cost-effective screening method. It is a qualitative, rapid test for the presumptive qualitative detection of E. coli O157 from a variety of food matrices, including meat and dairy products. This test is for point-of-care use or reference lab settings. The test is designed as an aid in the detection of E. coli O157 and should be used as an adjunct to culture. Features/Advantages of the E. Coli Test Kit Rapid strip test method (results; 20 minutes of incubation) High sensitivity (1X104 CFU) High reproducibility PRINCIPLE OF THE TEST The E. coli O157 Strip Test Kit is designed to detect E. coli O157 antigens in contaminated food samples. The test is a rapid qualitative lateral flow test that is based on the use of E. coli O157- specific antibodies. A reaction between the enriched positive sample and the colored particleconjugated antibody forms a complex that migrates along the membrane. If E. coli O157 is present in the sample, two red lines will form on the test cassette window. In the absence of E.

3 coli O157, only one red line will form at the control line (C), which is built in to assure that the test has been carried out correctly. KIT CONTENTS, STORAGE AND SHELF LIFE The E. coli O157 Test Strip Kit has the capacity for 25 determinations, not including up to 4 positive controls. The shelf life is 12 months when the kit is properly stored. Kits Content Amount Storage E.Coli 0157 Test Cassette 25 cassettes 4 0 C E.Coli 0157 positive control (500x stock) Resuspend tube in 0.5 ml PBS C SENSITIVITY (DETECTION LIMIT) The sensitivity limit is between 1X104 CFU - 1X105 CFU. REQUIRED MATERIALS NOT PROVIDED WITH THE KIT 37 C Incubator Blender or stomacher/stomacher bags Disposable plastic pipettes Disposable polypropylene tubes Distilled or deionized water EC medium, modified with novobiocin (for meat and meat products) TSB medium, modified with novobiocin (for dairy) Autoclave SAMPLE PREPARATION Add 25 g of food sample or 25 ml liquid sample with 225 ml EC medium, modified with novobiocin (for meat or meat products) or TSB medium, modified with novobiocin (for dairy products) and homogenize for 2 minutes in a Stomacher if needed. Mix gently and place in a 37 C incubator or water bath for hrs. Allow it to cool to room temperature. (The following steps are not mandatory for the test performance but can be done in order to reduce the potential risk of contamination when handling live bacteria).

4 Transfer approx. 1-2 ml of enrichment culture to an appropriate tube. Place tubes in boiling water bath for 15 minutes. Remove and allow sample to cool to room temperature E.COLI 0157 TEST STRIP KIT PROTOCOL Test Procedure Remove the E. coli O157 test cassette from the pouch and place on a flat surface. Using a micropipette and disposable pipette tip, add 100 μl of cooled enrichment sample (or the E. coli O157 Positive Control) onto the test cassette sample reservoir. Alternatively, using a disposable transfer pipette, squeeze the pipette bulb, insert stem into the cooled enrichment sample and release pressure on the bulb. Dispense five free falling drops into the sample port on the test device. Incubate at room temperature for 10 minutes. Analyze the test cassette within 20 minutes of incubation. Note: It is recommended to read results no later than 25 minutes after the sample application, before the device starts to dry. INTERPRETATION OF RESULTS 1. Negative results The test is negative if only one colored line appears in the C (control) area. 2. Positive results The test is positive if two colored lines appear. One colored line will appear in the T area (Test line) and one in the C (control) area. A colored line with any intensity in the T (Test line) area should be considered positive. Colored lines may be lighter or darker than each other. A highly positive result can produce an intense red line at the T area (Test line) and a faint line at the C (Control) area. This would still be considered a positive result. 3. Invalid results The test is invalid if no colored line appears in the C (control) area even if a colored line appears in the T (Test line) area. If no colored line appears in the C (control) window, add an additional broth mixture and wait an extra 4 minutes. If a colored line still does not appear in the C (control) window, the test is invalid and should be repeated using another device. Colored lines which appear after 20 minutes are not diagnostic and should be ignored.

5 WARNINGS AND PRECAUTIONS The National Food Safety Foundation strongly recommends that you read the following warnings and precautions. Periodically, optimizations and revisions are made to the kit and manual. Therefore, it is important to follow the protocol coming with the kit. E. coli 0157 isolates have been shown to be infective at very low dosage (less than 50 bacteria). Extreme care should be used in handling samples, enriched media and used test strips. Ensure all biohazerdous waste is disposed of appropriately. Decontaminate the test strip devices, enrichments, tubes and pipettes by autoclave in accordance with local, state, and federal regulations. Do not use the kit past the expiration date Incubations of assay should be timed as precisely as possible. Always store test strips in sealed bags with a desiccant to maintain stability. Interference caused by the use of alternative enrichment broths cannot be excluded ORDERING INFORMATION In case you want to place your order for this kit or you want more information about our Testing Kit products, you can reach us by calling / or visit our website; You can also us on foodsafety@foodsafetyltd.com or visit our office which is located in Kulambiro-Ntinda on Plot 1099, Block 215, Kondogolo Zone, Off Ntinda Kisaasi Road on Kulambiro Ring road near East High School Ntinda, Kampala- Uganda