Transia Plate Gluten - IUL Instruments GmbH

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Transia Plate Gluten Dilution of the liquid extract The recommended dilutions for different types of food are found in Table 1. In order to reduce the risk of matrix interference a sample needs to be diluted at least 1/50 before performing the assay. The standard curve will give an accurate reading within the range of the standards. If the sample has a concentration outside this range it will need to be diluted further. We recommend that you make at least two dilutions for each sample. For a semi-quantitative test and for unknown samples prepare the dilutions 1/50, 1/250 and 1/1,000. Note that this type of aqueous ethanol solution, such as the liquid extract, is very difficult to pipette accurately. After dispensing the extract into the dilution tube, rinse the tip several times up and down with the buffer in the dilution tube. Dilution 1/50 Add 100 µL of the extract to 4.9 mL of the dilution buffer 1X. Dilution 1/250 Add 1 mL of dilution 1/50 to 4 mL of the dilution buffer 1X. Dilution 1/1,000 Add 1 mL of dilution 1/250 to 3 mL of the dilution buffer 1X. Dilution 1/10,000 Add 1 mL of dilution 1/1,000 to 9 mL of the dilution buffer 1X. Dilution 1/20,000 Add 1 mL of dilution 1/10,000 to 1 mL of the dilution buffer 1X. TABLE 1 Sample Expected Dilution gluten content Starch quality control or “gluten-free” products e.g. dietary bread, baking mix, baby food, 1/50 milk drink 0-0.02% to 1/1,000 Soup, processed meat, analgesic, icing sugar 1/1,000 to mix, pet food 0.1-2.0% 1/10,000 Biscuits, flour mixes, bread crumb preparations, noodles, 1/10,000 pasta >2.0% and higher NOT COM 111E 11/00 4 (8)
Transia Plate Gluten Assay procedure Important: Allow the reagents to reach room temperature (18-25°C). Remove them from the box at least one hour before use. Shake the reagent vials before use. Do not interchange reagents between kits with different batch numbers. The washing step is very important. When washing, direct a strong beam at the bottom of the well. 1. Attach the required number of strips to the plate: two wells for the negative control and two wells for each dilution of sample extracts. For a semi-quantitative assay add two wells for each reference starch. For a quantitative assay add two wells for each standard. Note that only dilutions 3-8 are used as standards. Return the unused strips to the zip-lock bag containing dehydrating agent and close it tightly. Write the position of the controls, standards and samples on the work sheet (ENR COM 113). 2. Distribute 100 µL of the negative control (use dilution buffer), standards and/or reference starches and dilutions of sample extracts into the assigned detection wells. 3. Incubate at room temperature (18-25 o C) for 40 minutes. Note that the incubation starts once the last well has been filled. Prepare the washing buffer and the conjugate solution - see Preparation of reagents. 4. Empty the plate over a container and shake out the remaining contents by briskly flicking your wrist. Rinse each well, keep the washing buffer in the plate for 5-10 seconds and then remove it by inverting the plate over the container and shaking out the remaining contents. Repeat the washing four times. At the end of the washing, drain the plate on absorbent paper, tapping it firmly. 5. Add 100 µL of diluted conjugate to each well using a multipipette. Be careful not to touch the wells with the tip. Mix the contents of the wells using a gentle circular motion. 6. Incubate at room temperature (18-25 o C) for 40 minutes. Note that the incubation starts once the last well has been filled. Prepare the chromogen/substrate mixture - see Preparation of reagents. 7. Perform the washing five times as described in step 4. 8. Add 100 µL of the substrate/chromogen mixture to all the wells, preferably using a multipipette. 9. Incubate at room temperature (18-25 o C) for 30 minutes. Note that the incubation starts once the first well has been filled. Mix the contents of the wells using a gentle circular motion. 10. Add 50 µL of stop solution (vial 10) to each well, following the same order used when the substrate/chromogen mixture was added. Mix the contents of the wells thoroughly to ensure complete colour conversion. The blue turns to yellow. 11. Read the optical densities using a plate reader at 450 nm (blank on air). NOT COM 111E 11/00 5 (8)
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Transia Plate Gluten - IUL Instruments GmbH

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