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munchkinette

School Project- What Should I Test For Wheat Contamination?

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I'm a biology major, and I'm currently taking a PCR and DNA sequencing class. I decided to test a variety of foods for wheat DNA for my final class project. I'm not testing for other gluten grains simply because I don't have time for the scope of the class. (I think testing for all three would involve designing my own primers as well.) Most companies who test for contamination use ELISA (antibody) testing, but some studies have been trying PCR, which directly tests for a specific DNA sequence.

So are there any products labeled "gluten free" or "no gluten ingredients" that might be interesting? I'm definitely going to visit Trader Joe's, since I read a lot of people having trouble with their products. (I have too sometimes.) I'm also testing a couple Bob's Red Mill things. I don't remember which products have been repeat offenders since I've cut entire stores and types of products out of my diet.

I'm in the US, and I really only have time for about 20 samples. Dry ingredients, cookies/crackers, and some sauces would probably work best.


Gluten free since Feb 2006, Dairy and Soy free since 2009

Anemic off and on since 2003

Negative tTG Ab, IgA, Gliadin Ab IgA, wheat allergy (IgE) blood tests (Feb 2006)

Positive wheat allergy skin test(Apr 2006)and dietary response (Feb 2006)

Celiac grandmother (Dx in 1940s, "grew out of it")

Training for my first triathlon to support the Crohn's and Colitis Foundation of America.

~Amy

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I would pick stuff with as few chemicals added as possible, to give your pcr a better chance of working. Also, don't forget you'll need a positive control, so maybe pick corn, or tapioca, something that's found in all the products you test. For testing flours you might have to spike your flour sample with your positive.


"But then, in all honesty, if scientists don't play god, who will?"

- James Watson

My sources are unreliable, but their information is fascinating.

- Ashleigh Brilliant

Leap, and the net will appear.

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soy flour... maybe 1 regular brand, and 1 of Bob's Red Mill.

i know they found A LOT in soy flour recently which makes me so SAD cause i used to use this for choco chip cookies and the taste was fantastic...

?


1986- Elevated Speckled ANA/no Lupus.negative Sjorgens

2008- AntiGliadin IGA/IGg~ Negative,TTG IGA/IGg~ Weak Positive, Endomysial Antibody~ Positive, IGA Deficient.

no biopsy (insurance denied)

6/2010- Enterolab Gene Test:

HLA-DQB1 Allele 1 0302

HLA-DQB1 Allele 2 0302

HLADQ 3,3 (subtype 8,8)

7/2010- 100% Gluten Free

8/2010- DH

10/2010-Hypothyroid dx-> 12/2010 Hashimoto's dx + 1/11- Graves dx :(

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I'd try a couple brands of oatmeal if your primers are specific enough. People say they have problems with Amy's pizza so that would be interesting.

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I would pick stuff with as few chemicals added as possible, to give your pcr a better chance of working. Also, don't forget you'll need a positive control, so maybe pick corn, or tapioca, something that's found in all the products you test. For testing flours you might have to spike your flour sample with your positive.

I'm using regular wheat flour as my positive control to make sure the primers are working. So far I've done Maseca corn flour, BRM Rice Bran, and a granola cereal with oats. I've also got a TJ's curry sauce and Quaker oats. I am a little concerned that the curry sauce will have some kind of inhibitor in it with all those spices. I'm doing that one because my sister buys it all the time and it smells awesome.

The soy flours sound like good ideas. I'm debating how many "dedicated facility" products I should try. I'm thinking Rice Chex might be interesting, and it doesn't bother me.


Gluten free since Feb 2006, Dairy and Soy free since 2009

Anemic off and on since 2003

Negative tTG Ab, IgA, Gliadin Ab IgA, wheat allergy (IgE) blood tests (Feb 2006)

Positive wheat allergy skin test(Apr 2006)and dietary response (Feb 2006)

Celiac grandmother (Dx in 1940s, "grew out of it")

Training for my first triathlon to support the Crohn's and Colitis Foundation of America.

~Amy

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What is the sensitivity of your test? Bob's tests their products to 20 ppm, so unless you test is more sensitive than that all Bob's gluten free products should end up negative. Amy's tests to 5 ppm, so unless your test is more sensitive than that, Amy's gluten free products should test negative. If you contact companies they will often tell you to what sensitivity they test to and that can give you an idea of what items might be best to test. For a less sensitive test you might want to test things like Kraft products which didn't test last I knew. You could also test items from a local gluten free bakery and gluten free meals in restaurants.

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What is the sensitivity of your test? Bob's tests their products to 20 ppm, so unless you test is more sensitive than that all Bob's gluten free products should end up negative.

That's a really good question. Are you controlling the study with an ELISA? PCR is very sensitive. If you're not controlling with ELISA you may need to do something like grind some corn flour from whole kernels of corn or rice flour from rice that you sort by hand and wash well, and spike in known amounts of gluten to get an idea of the sensitivity. I believe flour is about 10% protein, so you'll get around 10 ppm gluten for each 100 ppm flour you add.

Here's a study on flour composition I found with Google.

http://www.aaccnet.org/cerealchemistry/articles/1999/0416-04R.pdf

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PCR tests for the presence of nucleic acid, so you can verify the presence of wheat with the proper controls, but can you quantify contamination? Can it detect 1 wheat kernel in a truckload of oats? Which gene are you testing for? Is it something expressed in the wheat seed or in the stem/leaf portion? Is it specific for a gluten protein gene? I think your results will be interesting, but it will not necessarily correlate with presence of gluten proteins, for which the ELISA will be more direct. Your test will tell only whether wheat was present in some form in the food you are analyzing, but not necessarily whether the contamination includes gluten or is significant in a dietary/allergen way. But I still think it's interesting and would like to see your results. Please follow up here!


Lee

I never liked bread anyway.....

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How about Food for Life brown rice tortillas? They had a recall in Canada on them recently for possible gluten contamination.


Proverbs 25:16 "Hast thou found honey? eat so much as is sufficient for thee, lest thou be filled therewith, and vomit it."

Job 30:27 My bowels boiled, and rested not: the days of affliction prevented me.

Thyroid cyst and nodules, Lactose / casein intolerant. Diet positive, gene test pos, symptoms confirmed by Dr-head. My current bad list is: gluten, dairy, sulfites, coffee (the devil's brew), tea, Bug's Bunnies carrots, garbanzo beans of pain, soy- no joy, terrible turnips, tomatoes, peppers, potatoes, eggplant, celery, strawberries, pistachios, and hard work. Have a good day! 🙂 Paul

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I'm planning on using real time pcr to quantify. I asked my professor about trying an ELISA test. He said this was way more sensitive because you can get down to the number of dna molecules, but that he didn't need ELISA for the purpose of the class project. I'm just learning these techniques, but his background is with plant genomics, including a few decades with corn.

I ended up picking a variety of different things- some are labeled as ELISA tested, some are made on shared equipment, some have no gluten ingredients and don't claim to be gluten free, and some are gluten free when packaged but I got them out of the bulk bins just to see what I'd get. They are all relatively homogeneous foods like flours, sauces, and crackers because I have to use small samples.

Here's an example of the kind of thing I'm trying to do. (My project is to learn techniques as they apply to certain types of research, not do original research.)

Previous Study


Gluten free since Feb 2006, Dairy and Soy free since 2009

Anemic off and on since 2003

Negative tTG Ab, IgA, Gliadin Ab IgA, wheat allergy (IgE) blood tests (Feb 2006)

Positive wheat allergy skin test(Apr 2006)and dietary response (Feb 2006)

Celiac grandmother (Dx in 1940s, "grew out of it")

Training for my first triathlon to support the Crohn's and Colitis Foundation of America.

~Amy

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That link was interesting. They found the PCR sensitivity to be somewhat lower than ELISA. Quantification looks pretty tricky compared with ELISA too, which is not terribly surprising. If you're not using ELISA at all, I'm really surprised your prof doesn't want you to spike a sample with a known amount of wheat flour. I'd require you to do that if I were grading. :P

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