More and more people have food allergies. Think of allergies to milk, nuts, egg or gluten. It is therefore very important for food producers to prevent cross-contamination with allergen-containing substances during production in food where these allergens do not occur. The article below clarifies which methods are suitable for testing for the presence of allergens. And above all, how to apply them in a smart and practical way.
Incorrect allergen labeling can have major consequences. It can lead to product recalls, fines from regulators such as the NVWA and FAVV or even worse health problems among allergic consumers. It is therefore very important to know whether or not the amount of allergens in the product is above the reference values (RfDs) in order to include the correct allergen information on the label.
Controlling allergens in food production requires focused controls on raw materials, surfaces, equipment and finished products. There are four commonly used testing methods, each with its own applications, advantages and limitations.
Rapid tests, such as test strips, are quick, simple and suitable for line clearance or raw material control. However, they are usually only qualitative and sensitive to disturbances caused by grease, pH or heating. They are convenient on the shop floor, but less reliable for complex products.
Surface swabs are used after cleaning to detect cross-contamination. They are effective for hygiene controls, but do not provide assurance about the final product. Results are highly dependent on proper execution.
Rinse water analyses are suitable for CIP systems and assess overall cleaning. They give an indication of residual allergens, but are dilute and less sensitive than product analyses.
Product analyses (via ELISA, PCR or LC-MS) provide the most certainty about the presence of allergens in the final product. They are accurate but time-consuming and sensitive to influences of processing or matrix effects.
In practice, a combination of methods is most effective: rapid tests and swabs for immediate verification and product analyses or rinse water for deeper verification. Provide good training, proper method selection and regular validation to keep allergen risks manageable.
A good test starts with a good sample. Whether you're taking a swab from a machine, testing a product or analyzing rinse water. How you do it determines the result.
Allergens can be detected in several ways. Some methods are explained below. Each method has advantages and disadvantages. Choosing the right technique depends on the product, processing and what exactly you want to measure.
The ELISA method is one of the most widely used techniques within the food industry for detecting allergens. A kind of "test kit" that recognizes proteins of allergens. Quickly and often very precisely. This method is based on immunological detection and is particularly suitable for detecting protein-based allergens such as milk, sesame, egg, peanut, soy, gluten, lupin, tree nuts, fish and shellfish.
ELISA uses specific antibodies that bind to an allergen protein. When this binding occurs, an enzymatic color reaction is initiated. The intensity of the color is proportional to the amount of allergen in the sample and is measured with a microtiter plate reader (spectrophotometer). Among other things, an incubator and pipettes are required to perform the analysis.
The method is not suitable for detecting allergens whose protein structures have been severely degraded, such as by heat treatment or fermentation. DNA-based allergens also cannot be detected by ELISA.
Performing an ELISA test involves a few steps at the laboratory:
A major advantage of ELISA is that commercial kits are available for a wide range of allergens. The method is also suitable for quantitative analysis, making it valuable for monitoring and regulatory compliance. These advantages are offset by some limitations. ELISA is sensitive to matrix effects: the composition of the food product (such as fat content, pH or processing) can complicate protein extraction, leading to underestimation of allergen concentration. In addition, cross-reactions may occur, where antibodies also react to related proteins that need not be harmful. Changes in protein structure, for example due to heating or enzymatic degradation, can reduce recognition by antibodies, leading to false-negative results. Furthermore, we know that ELISA mistakes peas for soy. And that fish, celery, mollusks and sulfite cannot be detected by ELISA.
Known from corona times; this technique looks for the DNA of allergens. Useful when proteins have been altered by heating. Unlike ELISA, PCR does not target proteins, but rather the genetic material of an allergen. This makes PCR particularly suitable for allergens such as crustaceans, fish, celery, mustard, lupin and mollusks. Also possible for soy, peanut, egg, tree nuts, sesame and as long as sufficient DNA is still present in the product.
PCR works by amplifying (multiplying) specific DNA fragments using primers that target characteristic DNA sequences of an allergen. During the process, these fragments are multiplied in a thermocycler. In real-time PCR, this process is monitored via fluorescence, which provides an indication of the amount of DNA in the sample.
This method is particularly useful in products where proteins may have been degraded or modified, but where DNA is still present. However, PCR is not suitable for products that have been processed such that the DNA has been completely degraded or removed, such as highly refined oils.
This test is only for producers with a fully equipped molecular biology lab and trained personnel.
Currently mainly a qualitative result. Quantitative determination is possible with a conversion factor. This establishes the link between the DNA and protein.
PCR is highly sensitive and able to detect low amounts of allergen DNA even in complex or highly processed products. It is also less dependent on the integrity of protein structures. However, PCR does not provide information on the presence of functional or intact proteins - and thus not directly on allergenicity. The allergens egg, milk and sulfite cannot be detected by real-time PCR.
The method requires specialized equipment and experience. In some cases, components of the food may inhibit PCR reactions (so-called PCR inhibitors), making detection less reliable.
LC-MS is an advanced analytical technique for identifying and quantifying specific peptide fragments of allergenic proteins. This method is broadly applicable to common allergens such as milk, egg, peanut, tree nuts, gluten and soy.
In LC-MS, proteins are extracted from the sample and then enzymatically degraded into small peptide fragments (often via trypsin digestion). These peptides are separated via liquid chromatography (LC) and then analyzed in a mass spectrometer. The combination of the mass and charge of the peptides makes it possible to detect specific fragments of allergenic proteins.
This method is very suitable for analyzing products containing allergens because the technique can also recognize partially degraded proteins.
This test can only be performed by external specialized laboratories.
LC-MS offers exceptional specificity, minimizing the risk of cross-reactions. It is also possible to detect multiple allergens in a single analysis (multiplex). In addition, LC-MS can detect partially degraded or processed proteins, which ELISA does not always succeed in doing.
These advantages are offset by higher costs and technical complexity. The method requires specialized equipment and expertise. Matrix effects - such as fats and salts - can also affect the ionization of peptides, which can reduce the accuracy of the analysis.
Flow cytometry is used less frequently in the food industry, but is gaining popularity because of its multiplex detection capability. This technique uses microscopic beads (microspheres) coated with specific antibodies directed against allergenic proteins.
When a food sample is mixed with these microspheres, allergens present bind to the spheres. A second fluorescent-labeled antibody is then added. The amount of fluorescence is measured in a flow cytometer, with each microsphere analyzed individually.
This method is suitable for protein-based allergens and offers similar applications to ELISA, but with the ability to measure multiple allergens simultaneously.
These tests are used only by large companies with advanced R&D facilities.
A major advantage of flow cytometry is its ability to multiplex: multiple allergens can be detected simultaneously in a single analysis. The method is more sensitive than standard ELISA and offers more flexibility for complex matrices.
The disadvantages lie mainly in cost and complexity. Flow cytometers are expensive and require specialized training. In addition, this method is less common in routine food industry practice.
Although less sensitive to some matrix effects than color reactions (as in ELISA), certain food components can still affect binding or fluorescence, particularly fats or acidic pHs. Again, processed or denatured proteins may affect recognition.
Sometimes tests give false results: they say it is an allergen when it is not (false-positive), or conversely that it is not an allergen when it is present (false-negative). Causes may lie in processing of the product, incorrect test kits or interfering substances in the sample. Therefore, it is important not to assume 1 test result but to repeat it at least 2x.
Reliable laboratories work according to strict quality rules (e.g. ISO 17025). This means that their tests are validated, personnel are well trained and results are reliable. Normec Group laboratories work under accreditation and even when analyses are outsourced, Normec Foodcare works exclusively with these types of laboratories.
Which method is best to choose per type of allergen. See below the advice** from the lab.
** Please note that this is an indication; this advice is based on the info available at the date of publication (July 2025) of this article. Always seek advice from the lab before scheduling and performing an analysis.
Normec Foodcare is all about food safety, quality and practical advice. Whether you want to know which test method to use, how to make your processes safer or how to comply with legislation. The experts at Normec Foodcare think along with you.
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