Meeting Your GLUTEN Testing Needs

Transcription

1 Meeting Your GLUTEN Testing Needs COELIAC DISEASE: True gluten intolerance or Coeliac Disease (CD) affects up to 1% of the EU, US and Australian populations but is largely undiagnosed. CD is caused when toxic proteins in cereal gluten damage the absorptive areas of the small intestine. CD manifests itself as e.g. intestinal pain, bloating and cramps and in children, as a failure to thrive. CD can only be reversed by strict adherence to a diet low in gluten. Increasingly many people also perceive themselves to be food intolerant and avoid wheat/gluten as a result. These phenomena have led to a rise in the availability of gluten-free food products, defined in Codex guidelines/eu Legislation as containing <20PPM gluten. Cereal derived foods with levels >20<100PPM are to be defined as very low gluten. A comprehensive range of RAPID on-site tests, quantitative Gluten-Check ELISA/Sample Extraction kits and Assayed Quality Control sets to assist in the accurate & cost-effective determination of low levels of cereal GLUTEN in food raw materials/commodities, environmental swabs, unprocessed/processed foods etc. GLUTEN Flow-Through (GFT) Tests: fast and sensitive tests for the presence of cereal gluten in food raw materials, finished products, surface/environmental swabs etc. The tests are designed for use in food factories with limited testing facilities or small laboratories with lower testing needs. Several samples can be tested within 20 minutes and gluten levels of parts per million (PPM; mg/kg) detected in food (swab samples <0.5PPM). GLUTEN-Check ELISA KITS*: designed for the measurement of cereal gluten within a nominal analytical range of PPM or more. They use mouse monoclonal antibodies to detect the omega gliadin fraction of wheat (Limit Of Detection <0.5PPM) as a marker of total gluten content. The kits are available with or without Extraction Kits see below, and in 48-, 96- & 192-well sizes. * Originally developed by Skerritt & Hill (JAOAC 74, 2, 1991, p ). GLUTEN EXTRACTION KITS: each contains 24 pre-filled tubes of Imutest s Extraction solution for gluten (which contains Tannin Binding/Protein Additive) with yellow dye as a pipetting aid. Homogeneous Laboratory Sample is weighed directly into the tube and extracted; the tubes can then be centrifuged and the extract diluted for ELISA testing; diluted extracts are pale yellow in colour to aid visibility when adding to microwells. GLUTEN ASSAYED QC SETS: 4 x 6g pots. Two pots in each set contain very low levels of gluten; two contain assayed amounts of gluten. These QC sets are designed to help verify the accuracy of whichever gluten analysis method you use. Diagnostic Innovations Limited Spectrum House, Llys Edmund Prys, St. Asaph Business Park, LL17 0JA, U.K. Tel: +44(0) Fax: +44 (0) food@imutest.com

2 SCHEMATIC DIAGRAM OF Imutest GLUTEN FlowThrough/ELISA METHODS: 1. Antibody coated 2. Gliadin ( ) in 3. HRP enzyme-labelled 4. Addition of TMB substrate, plastic micro-well ethanolic food antibody binds in turn which develops blue or test area on extract binds to to gliadin bound to colour in presence of GFT card. antibody. antibody on well*. HRP enzyme*. *In GFT Tests, enzyme label is replaced with an intensely coloured gold particle which is directly visible after testing. Fast, sensitive test for cereal GLUTEN GLUTEN TEST Use for food samples/surface swabs For testing on-site or in small labs EXTRACTION: Food samples are extracted with the same highly effective solution used in GLUTEN-CHECK ELISA kits, provided in pre-filled tubes. The mixture is shaken to extract gluten and the contents allowed to settle/centrifuged to clarify the extract. DILUTION: Seven drops of the food extract are added to a pre-filled tube of Diluent Liquid and mixed. NOTE: The sample extract can be retained for later ELISA testing if required. PERFORMING the TEST: Diluted extract is added to the Gluten Flow-Through card. The extract slowly (3-5 minutes) percolates into the card through the read-out area and any gluten present binds to the Test spot. The well and read-out area are wiped with an absorbent pad. Finally, a pre-filled gold visualisation reagent is added to the read-out area; as it flows into the device (3-5 minutes) the reagent links to any gluten present on the Test spot, causing a pink/red spot to appear. The darker the Test spot the more gluten is present in the food being tested. The GLUTEN Flow-Through test result is then read visually see next page. Page 2 of 8

3 READING THE GFT TEST: The appearance of a pink/red spot on the left of the test area indicates the presence of gluten at a level of 10-20ppm or more; the darker the Test spot the more gluten in the food. Since the test is sequential in nature it cannot be overloaded with gluten. A pink/red control spot of medium intensity should always appear on the right hand side of the read-out area as an indication that the food extract is suitable, the test has been performed correctly and all reagents are functioning correctly: Above (left to right): Negative (bread) <5ppm; Low Positive (biscuit) 20ppm; Low Positive (bread) 40ppm; Medium Positive (2 drops of 300ppm wheat AOAC B starch extract); High Positive (low gluten wheat starch). Left Spot: TEST (GLUTEN) Right Spot: PROCEDURAL CONTROL Each GLUTEN Flow-Through Test pack (right) comprises: Tube containing 3.5mL of (yellow) Extraction Solution. Two small spoons for adding sample to Extraction Solution. Centrifugation tube (not used if sample allowed to settle). Two pipettes for removing fat layer from and/or diluting extract. Tube containing colourless Diluent Liquid. Gluten Flow-Through Device (in a protective pouch). Absorbent pad for wiping/drying test area. Tube containing (pink) anti-gluten/gold particle Visualisation Reagent. What else do you need to perform the GLUTEN Flow-Through Test? Sample blender to ensure food sample homogeneity (not required for e.g. flours/fine powders/liquids) Mini centrifuge (Optional: to speed up clarification of the sample extract). Balance (0.02g or 0.01g display) for accurate weighing of food samples (Optional: sample size can be estimated using the spoons provided, but these allow only a rough approximation). Page 3 of 8

4 GLUTEN-CHECK TM ELISA SAMPLE PREPARATION Prepare Sample by grinding/chopping/blending until homogeneous. Add 1 part Test Portion to 9 parts Extraction Solution (1/10). Extract Mix. Separate Remove Dilute Shake 2 mins; 45 C with mixing. Shake 2 mins; 10 mins centrifuge OR 30 mins settle. a portion of the extract above the food pellet. Sample/Control extracts in Diluent Solution 1 (1x). ELISA PROTOCOL: Pipette Mix. Incubate 100uL diluted Sample/Control Extracts & Standards into wells. at room temperature for 40 minutes. Wash wells three times with wash solution 1X. Pipette Mix. Incubate 100uL Anti-Gliadin HRP into wells. at room temperature for 40 minutes. Wash wells four times with wash solution 1X. Pipette Mix. Incubate Pipette Mix. Read Calculate 100uL TMB Substrate reagent into wells. at room temperature for 15 minutes. 100uL Acid Stop Solution 1 into wells. wells at 450nm wavelength. mg/kg (PPM) Gluten results for all Controls/Samples. The figure (right) shows an example Standard Curve obtained using the Imutest Gluten-Check ELISA method. At present there is no agreed calibrator to help support gluten analysis. The Imutest Gluten-Check ELISA has been calibrated using samples previously characterised in the FAPAS Proficiency Testing Scheme ( and D.I.L. s own in-house Q.C. panel of gluten free/low gluten food samples. Page 4 of 8

5 Swabbing for GLUTEN on Environmental Surfaces SWABBING TECHNIQUES can help validate and verify clean-down routines of food suppliers/ manufacturers, catering establishments and analytical laboratories. Imutest Environmental/ Surface Swabbing kits have been designed to make the process simple, effective and safe. Imutest-ESS Kits contain: 25/100 individually pouched sterile flocked nylon swabs. 25/100 pre-filled tubes of stabilising/preservative swab solution. Instructions For Use; swabbing Identification & Results Form. Features of the kits: Pre-filled swabbing solution Blue dye added NON-TOXIC preservative/stabiliser Benefits to you: Accurate volume; less hands-on time Highly visible; aids removal No risk of food contamination How does the Imutest-ESS system work? Swabs are wetted with swabbing solution and used to swab a representative area of surface, equipment, air filter etc. The swab is replaced in the tube, the end snapped off and the tube capped securely. Gluten in the solution is stabilised for several days, even at elevated room temperatures. The swab solution can be used to carry out a GLUTEN FlowThrough test immediately or sent to Diagnostic Innovations Limited (D.I.L. see contact details on page 8.) or your preferred laboratory for ELISA testing. Imutest Gluten FlowThrough Swabbing Kits Specially packed combinations of swabs, ESS swabbing solutions, GFT test cards and visualisation reagent tubes all you need for on-site testing of swab samples for gluten content. These kits enable swab samples to be tested for gluten (limit of detection approximately 0.5PPM [mg/kg]) virtually anywhere in ten minutes or less. Page 5 of 8

6 Gluten Testing: Contract Analysis at D.I.L. D.I.L. offers contract analytical services to test the gluten content of your: Raw materials. Part-processed foods. Finished Products Environmental swabs All analyses are performed using our High Sensitivity Gluten-Check ELISA method. You can choose to receive Qualitative ( Yes/No against a specified PPM threshold) or fully Quantitative (PPM values). Testing is normally performed in duplicate against either a Negative & Positive Standard (Qualitative testing) or a full Standard Curve (5-110 PPM [mg/kg] Quantitative testing); at least one Procedural Q.C. Control is also tested at the same time. Indicative Contract Analysis prices (please ask for a specific price quotation for your testing requirements): No. of samples sent Qualitative Testing (Price per sample tested*) Quantitative Testing (Price per sample tested*) 2 food samples food samples food samples food samples swab samples swab samples swab samples swab samples 9 17 * Normally 3-4 day turnaround; costs may increase for shorter, or decrease for longer times to results. Page 6 of 8

7 PRODUCT INFORMATION (1) GLUTEN TESTING PRODUCTS Imutest GLUTEN-CHECK ELISA ANALYSIS PRODUCTS Gluten-Check ELISA (48 wells + 1 Extraction kit 24 tubes) Gluten-Check ELISA (96 wells + 2 Extraction kits 48 tubes) Gluten-Check ELISA (192 wells + 4 Extraction kits 96 tubes) R6005-1E R6006-2E R6007-4E Wheat omega gliadin (Skerritt) ELISA Including extraction kits (LOD <0.5ppm; range 5-110ppm gluten) Gluten Extraction Kit (24 tubes) A ml/tube of gluten extraction solution with yellow dye; suitable for all samples, including those with high tannin Gluten-Check ELISA (48 wells) R6005 Gluten-Check ELISA (96 wells) R6006 Wheat omega gliadin (Skerritt) ELISA (LOD <0.5ppm; range 5-110ppm gluten) Gluten-Check ELISA (192 wells) R6007 Imutest GLUTEN FlowThrough RAPID TESTING PRODUCTS (box of 20 tests - no plastic parts) A6021 (box of 50 tests) (box of 10 tests) (box of 5 tests) A6028 A6013 A6011 Wheat omega gliadin (Skerritt) FlowThrough rapid card tests (LOD 20ppm) (envelope with 2 tests) A6012 RAPID TESTING ANCILLARY EQUIPMENT (OPTIONAL) Mini centrifuge MINICENT 6-position mini centrifuge (200 rpm) 1-2 weeks Mini balance MINIBAL 60g mini balance (weighs to 0.01g) 1-2 weeks Imutest ENVIRONMENTAL/ANALYTICAL SWABBING Test Kits Environmental Surface Swabbing Kit (100 swabs & tubes) Environmental Surface Swabbing Kit (25 swabs & tubes) A6008 A6009 To assist with the validation of cleaning regimes and verification of clean-down in manufacturing & analytical environments Imutest SWABBING + RAPID GLUTEN PRODUCTS GFT Swabbing Kit (box of 20 sets) GFT Swabbing Kit (box of 10 sets) GFT Swabbing Kit (box of 5 sets) A6010 A6019 A6020 E.S.S. Swabbing Sets + GLUTEN FlowThrough rapid card tests Wheat omega gliadin (Skerritt) (LOD 1ppm) For testing environmental swabs for gluten "on site" in 10 minutes or less Page 7 of 8

8 PRODUCT INFORMATION (2) ALLERGEN-CHECK ELISA KITS Product Name Wells Cat. No. Product Description 192 R6011-4E Fish protein detection by ELISA Fish-Check 96 R6010-2E (LOD <1ppm; range 5-125ppm cod) Including Extraction Kit 48 R6009-1E INCLUDING 1 x A6015 for each 48 wells. Fish-Check 192 R6011 Fish protein detection by ELISA 96 R6010 (LOD <1ppm; range 5-125ppm cod) 48 R6009 Almond-Check* 96 R6018 Almond protein detection by ELISA 48 R6019 (LOD <0.25ppm almond; range ppm almond) β-lg/milk-check* 96 R6036 Beta-Lactoglobulin (milk/whey) detection by ELISA 48 R6037 (LOD <0.1ppm Skim Milk Powder; range 0.2-7ppm) Casein/Milk-Check 96 R6022 Casein (milk/caseins/caseinates) detection by ELISA 48 R6023 (LOD <0.2ppm Skim Milk Powder; range ppm) Crustacea-Check 96 R6026 Crustacea protein detection by ELISA 48 R6027 (LOD 1ppb tropomyosin; range ppb) Egg-Check* 96 R6032 Egg White protein detection by ELISA 48 R6033 (LOD <0.2ppm Egg White Protein; range 0.4-8ppm) Hazelnut-Check* 96 R6014 Hazelnut protein detection by ELISA 48 R6015 (LOD <0.5ppm hazelnut; range 1-40ppm hazelnut) Lupin-Check* 96 R6030 Lupin protein detection by ELISA 48 R6031 (LOD <0.5ppm lupin; range 2-30ppm lupin) Mustard-Check* 96 R6012 Mustard protein detection by ELISA 48 R6013 (LOD 1ppm mustard; range 2-50ppm mustard) Peanut-Check* 96 R6020 Peanut protein detection by ELISA 48 R6021 (LOD <0.5ppm peanut; range 1-30ppm peanut) Sesame-Check* 96 R6028 Sesame protein detection by ELISA 48 R6029 (LOD 0.2ppm sesame; range 2-20ppm sesame) Soya-Check* 96 R6024 Soya Trypsin Inhibitor detection by ELISA 48 R6025 (LOD <1ppm soya; range 2-25ppm soya) Walnut-Check* 96 R6016 Walnut protein detection by ELISA 48 R6017 (LOD <1ppm walnut; range 2-50ppm walnut) *Food sample extracts from these kits are fully compatible and can be used in any other of the kits marked *. NOTE: The LOD & Range data given above refer to PPM of the commodity itself and - unless indicated otherwise - NOT PPM of allergenic protein, for which the equivalent values are substantially lower. Diagnostic Innovations Limited 2011 LA012 Flyer_Gluten Testing_v02 Page 8 of 8