FAQs

IN-HOUSE ANALYTICAL TESTING

How do I send in samples?

Sample should be placed in a well-sealed container: such as a Whirl-Pak®, Ziplock®, or plastic container with a lid. Liquids can be sent in their original commercial packaging (with tight-fitting lid) or in plastic bottle. Perishables should be accompanied by an ice pack (not ice in a bag). For multiple samples, ensure that individual packages/containers are sealed to prevent cross-contamination during shipping. Also include a completed Sample Request Form. The information provided on the Sample Request Form will be used exactly as supplied for your test report, so fill out all information accordingly (i.e., name, contact info, sample ID, etc.).

How much of a sample should I send?

A minimum of 100 grams of product is requested when sending in a sample for testing. We retain a portion of the sample for three months before being properly destroyed; we do not retain swab samples. If you do not have 100 grams, we can manage with 20 grams in most cases.

What if I want my sample returned to me?

If you want your sample returned, you must state so on the Sample Request Form. You will be charged for shipping.

When can I access my test results?

Testing in our laboratory takes up to three (3) business days after the sample is received in our laboratory. Special testing requests for drugs, hormones, toxins, or antibiotics may take up to ten (10) days depending on kit availability. Following testing, the results are reviewed for quality assurance. After review and confirmation of successful payment, the test report is made available on our website and can be mailed to the address provided if requested. Results can only be provided to the mailing and email address supplied on the Sample Request Form.

How do I access my test results?

After your test has been completed and the results have been approved for release, test reports are posted on our secure server as pdfs; a hard copy is sent by mail if requested. The hard copy of the test report(s) can be sent only to the mailing address provided on the Sample Request Form. You will receive an email only to the single email address provided on your Sample Request Form notifying you that the test reports are ready. If you are a new customer, you will also receive a preceding email with your user information (login and password). If multiple users will need access to the test reports, we recommend using an email address that is accessible to all necessary individuals.

 

PAYMENT INFORMATION

How do I pay for testing kits or services?

Payments are normally made by credit card or wire transfer; we also accept checks. However, your products will be shipped after the check has cleared. Test reports are released following confirmation of payment. Net-30 accounts can be established upon request and require credit references.

 

SPECIATION TEST KITS

How sensitive are the kits?

You can find specifics listed on the kit data sheets.

Are there limitations to their use?

These assays are intended for use as qualitative tests to determine the presence of meat species content. The color development is proportional to the original amount of specific antigen in the extract, but these assays are not designed to quantify the amount of species tissue in a sample. Variations in the sample content (e.g., % lean tissue, % moisture, % fat, etc.) and variations in sample treatment (e.g., cooking times, temperatures, etc.) of samples will affect the amount of detectable antigen in the extract. Therefore, the level of the antigen present and the intensity of the color reaction are influenced by the sample composition, processing, and other factors.

Can I use an ELISA-TEK® Cooked or Raw Meat Speciation Kit to detect the presence of chicken egg proteins in my product?

For raw (uncooked) products, the Raw Meat Poultry ELISA kit will react strongly in the presence of raw whole egg, egg yolk and egg white with yolk contamination. These materials, when extracted as per the kit instructions, produce signals which are similar to a 100% chicken tissue positive control. Raw egg white alone will give a signal greater than a 1.0% chicken tissue positive control.

For cooked products, cooked whole egg, egg white, egg yolk and yolk-contaminated egg white all give negative signals in the Cooked Meat Poultry ELISA when extracted as per the kit instructions; therefore, processed dried egg white additive should produce similar results.

How long does testing take and how many samples can be tested?

Excluding sample preparation times, the Raw Meat Species and the Cooked Meat Species tests take approximately one (1) and three (3) hours, respectively. Both kits can be used as a 96-well unit or may be divided into a variety of strip formats depending on the number of samples to be analyzed and the number of replicates desired. USDA-FSIS protocols require use of quadruplicate microwells for presumptive positive samples, although duplicate microwells for each control and sample extract are acceptable for screening purposes.

What is the difference between the raw and cooked kits?

The ELISA-TEK® Raw Meat Species Kits use antibodies raised to species-specific serum proteins, whereas the ELISA-TEK® Cooked Meat Species Kits utilize antibodies raised to heat-resistant glycoproteins found in muscle and other blood-fed tissues.

How are results determined? Do I need special equipment?

The completed assay may be assessed visually or, more accurately, with the aid of a microplate reader or spectrophotometer. Visual assessment of the presence of green coloration can be aided through the use of appropriate positive controls representing suitable ‘cut-off’ levels (e.g., 1% tissue).

Microplate readers should be able to read absorbances at 414 nm (405-420 nm acceptable) and 492 nm (485-500 nm acceptable) for the Cooked Meat Species, 450 and 630 nm for Raw Meat Species, or 450 nm for MELISA-TEK assays. Test samples are generally classified positive or negative based on the relation of their mean absorbances to the absorbance of a 1% positive tissue control. If following USDA-FSIS protocols using a dual wavelength reader, read the absorbance at 414 nm with a 492 nm reference filter (485-500 nm acceptable). Specific USDA assay validity parameters are provided in the kits’ Instructions for Use. Sample data entry sheets (in .xls format) are available from the product pages of the website to assist in data analysis (coming soon).

What are the potential causes of erroneous results?

Poor reproducibility of replicate samples or significant color in the Negative Control wells (high non-specific binding values) are often the results of insufficient washing or can be attributed to contamination of the TMB or ABTS substrate or splashing of the conjugate.

Cross-contamination of tissue extracts can also lead to false positive results. This can be controlled during the sample preparation stages by using disposable materials and thoroughly cleaning equipment between stages to remove all traces of possible contamination. Wherever possible, all reusable equipment should be very easy to decontaminate, since both serum albumin and cooked protein antigen can be very difficult to clean from surfaces.

What is the difference between the raw albumin control and the lean tissue control in the Raw Meat Speciation Kits? Which should be used?

The raw controls can be used as true positive and negative controls in a raw test but are NOT equivalent to 100% tissue positive controls and should NOT be used for preparation of 1% tissue positive controls. Note that care should be taken not to cross-contaminate meats used for preparation of tissue controls. Tissue control extracts are more representative of ‘real world’ samples (e.g., a complex tissue matrix that is similar to a sample meat extract) than pure control, and we recommend that a 100% tissue control be used for regulatory protocols.

 

ALLER-TEK™ GLUTEN ELISA KIT (96 wells)

How sensitive is the kit?

The ALLER-TEK™ Gluten ELISA kit is designed with a range of detection from 5-80 ppm. Using dilutions of up to 1,000× with sample dilution buffer, the upper limit of detection can be extended to 80,000 ppm.

Are there limitations to their use?

This assay is designed to accurately quantify gluten content in food and beverage products and is approved as a Performance Tested MethodSM (Cert. #081202). The antibody in this assay recognizes multiple fractions of gluten, including both glutenins and gliadins, and this assay has been calibrated for the quantification of total wheat gluten; the detection of barley gluten can be determined by using our barley standards (Product #510821I).

How long does testing take and how many samples can be tested?

Sample extraction takes as little as 20 minutes with centrifugation or 50 minutes without. The remainder of the assay can be completed in approximately two and a half hours. The assay comes in a 96-well format; in addition to the standards, positive control, and negative control, up to 26 samples may be run in triplicate.

How are results determined? Do I need special equipment?

The completed assay is measured with the aid of a microplate reader or spectrophotometer by determination of absorbance at 450 nm.

Results are determined by plotting the standards on a graph and comparing the mean values of test samples to the standard curve. For assistance in plotting standards and calculating values for samples, please refer to the sample data entry form on the product page (coming soon).

What are the potential causes of erroneous results?

Poor reproducibility of standard or replicate samples or significant color in the Negative Control wells (high non-specific binding values) are often the results of insufficient washing or can be attributed to contamination of the TMB substrate or splashing of the conjugate.

Cross-contamination of samples can also lead to false positive results. This can be controlled during the sample preparation stages by using disposable materials and thoroughly cleaning equipment between stages to remove all traces of possible contamination.

Why does your test provide different results than a comparable assay?

There are several antibodies used for gluten detection in food products; some of the most common are 401.21 (Skerritt), R5 (Mendez), and G12, with other mono- and poly-clonal antibodies also in  use. These antibodies have been rigorously tested in interlaboratory studies around the world. Our ALLER-TEK Gluten ELISA and EZ Gluten lateral flow device use the Skerritt antibody, although it is important to use the right antibody for each test. Different antibodies are better suited for different testing whether the goal is accurately measuring total gluten content, specific measurements of gliadin from certain grains, detection of hydrolyzed gliadin, or identifying specific components that may be harmful to patients with Celiac disease.

The Skerritt antibody has been in use for over 30 years and is found in ALLER-TEK, EZ Gluten, Veratox and other assays and was selected to recognize ω-gliadin, but recognizes sequences in both gliadins and glutenins, providing the broadest view of gluten content. This antibody appears to detect gluten similarly across all gluten-containing grains, with slightly reduced sensitivity to barley gluten in some applications. Therefore, a barley-specific standard curve is necessary when specifically measuring barley gluten. This antibody is useful for analyzing raw or processed foods, beverages, and surfaces when total gluten content is of concern.

The R5 antibody has been in use since 2003 and is found in RIDASCREEN and Veratox assays. It was developed in 2003 to recognize the QQPFP sequence in rye gliadin (secalin). This sequence is also found in prolamins from wheat and barley, and is capable of providing measurements more specific to the antigenic portion of the wheat α-gliadin protein. Shortly after its release, the R5 was adopted as the preferred gluten antibody in the Codex Alimentarius of the WHO in 2008, putting it at the forefront of gluten testing. The R5 antibody has been used in competitive assays, which are most useful when gluten has been broken down or degraded, such as through hydrolysis or fermentation, making this antibody ideal for testing hydrolyzed or fermented products.

The G12 antibody is the newest of the most common antibodies and is found in AgraStrip, AgraQuant, and GlutenTox assays. It recognizes the 33-mer (LQLQPFPQPQLPYPQPQLPYPQPQLPYPQPQPF) of α-gliadin, one of the peptides which triggers an immune response in T-cells from patients with Celiac disease. This antibody is specific to this particular immunostimulatory sequence of gliadin and is useful for analyzing products when Celiac disease is of particular concern.

These are not the only gluten detecting antibodies in use with some kits using polyclonal antibodies or antibody cocktails to ensure gluten detection across strains and varieties of grains. For general detection of gluten, a broad-specificity antibody, polyclonal antibody, or antibody cocktail may be preferred, while a highly specific antibody may be necessary for the measurement of a specific epitope. Particularly with lateral flow devices, some are made for ease of use for home users, while others are more robust and versatile, suitable for use in industry. For these reasons, the purpose of testing must be considered when determining whether a particular test will meet your needs.

The antibodies and potential applications of each assay are often available upon request or on the manufacturer’s website.

EZ GLUTEN™ LATERAL FLOW KIT

What if my sample absorbs all of the extraction solution?

If a sample absorbs all of the extraction solution, and there is no clear liquid layer on top of the sample, try letting the sample settle for an extra 5 minutes (10 minutes total). If this does not result in enough clear liquid to perform the test, then it may be necessary to repeat the test with a new vial of extraction solution, this time using only half a spoonful of the sample. Note that using less sample will decrease the sensitivity of the test, but may be necessary for highly absorbent samples.

As an ingredients manufacturer, are there any special considerations when testing raw ingredients in my lab?

We have successfully tested over 100 products and ingredients as part of the EZ Gluten validation. Ingredients which are strong dyes, acids, or bases may interfere with assay performance. Specifically, deep red dyes may make the test strip difficult to read. The assay is also functions best in a pH range of 6-8; the extraction solution is able to buffer most samples, although strong acids or bases may require neutralization with HCl or NaOH after extraction to bring the final pH into range.

What is the Hook line?

Some lateral flow tests do not function in the presence of very high levels of target material. This false negative result is referred to as a “hook effect”. To allow for testing of samples containing very high levels of gluten, EZ Gluten™ test strips include a hook line. Under very high gluten conditions, unbound gluten can block the test line, interfering with binding of the antibody-bound antigen, resulting in a false negative result. The hook line found on the EZ Gluten™ test strip allows the user to determine if a weak or absent signal at the test line is due to low levels of gluten, or to excessively high levels of gluten. If the Test line is weak or negative, and the Hook line is present, then the weak or negative signal is due to low levels of gluten.

– If the Test line is weak or negative, and the Hook line is not present, this indicates a hook effect, or a high level of gluten in the sample. The EZ Gluten™ test can detect gluten levels as high as 100,000 ppm (10%).