The allergic reaction is mediated by IgE antibodies (WHO/IUIS 1984). These molecules are present in the blood and can bind to a specific allergen. There are a lot of different IgE to match the diversity of existing allergens and each person possesses a different range of IgE. When a patient is exposed to an allergen, this allergen enters the blood vessel and binds to an IgE. This activated IgE will then bind a receptor of the mast cell (an immune cell in the blood). When the mast cell is activated by the IgE, it will release a histamine molecule. Histamine will then trigger the allergic reaction. The blood vessel dilates, which causes visible swelling and redness or even anaphylaxis in the worst case. If a person does not possess this same IgE in their blood, they cannot be allergic to these molecules. That is why the detection of IgE is an usual method to help diagnose allergies.

It is mandatory to identify the cause of the allergy. There are two main methods to do so: the prick test and the blood test.

The prick test

The Prick test consists in applying purified allergens under the patient's skin. After 15 minutes, the skin is observed. If there is redness and swelling around the exposure areas, it means that an allergic reaction was triggered by the chosen allergen. It is usually performed on the arm or the back of the patient and could be unpleasant and dangerous in case of severe allergy. A dozen different allergens could be assessed. The results could vary a lot, depending on the state of the immune system or medication and often presents false-positives (Mayo Clinic). Furthermore, the number of allergens tested are limited, no more than a few dozen per test. In practice, only the most common allergies are tested. However, there are a lot of different allergens for the same cause. For example, there are 18 peanut allergens (WHO/IUIS 1984), but ara h 2 is the most tested one. Consequently, some patients are declared not allergic to peanuts while they react to other peanut allergens than ara h 2.

During our Human Practice process, allergists have mentioned that the exposure under the skin triggers a higher immune reaction than ingestion or exposure through airways.

The blood test

Blood tests are usually performed using ELISA or Immunocap methods. They all consist in measuring the concentration of an antibody recognizing a specific allergen in the patient serum. Purified allergens are immobilized at the bottom of a well or a biochip in which the serum is added. The patient’s IgE binds to the allergens and stays in the well or on the biochip even after washes. A second labeled anti-IgE antibody is then added so that IgE presence is detected. If there is a signal, it means the patient has a specific IgE to the target allergen in their blood and is probably allergic to it (Rame and al. 2005). Furthermore, there are false negatives and the number of allergens are limited (about 10 for an ELISA test, and a hundred for the Immunocap test). Even if data about false positives is not available, allergists contacted for our Human Practice strategy also affirmed there are false positive results, which means some patients have positive results during their test even if they are not allergic to the targeted allergen. It mostly occurs with biochips, especially when there are a lot of allergens tested (more than 50)(Menardo, personal communication, July 07, 2022).

For allergies, the best way to avoid danger is to avoid allergens. Therefore, it is essential to be perfectly diagnosed. However, current methods of diagnosis don’t test every allergen. That is why it is important to overcome limitations of current allergies tests so there are no more undiagnosed allergies. The iGEM INSA-UPS Toulouse have chosen during this year to overcome the limitations of classical allergic tests by inventing a new and innovative way to detect IgE, using synthetic biology. Our goal is to detect the presence of any IgE responsible for an allergy. Therefore we aim to engineer two bacteria able to aggregate when, and only when, a specific IgE is present. The first strain, called strain “A”, displays one type of allergen on its surface, which binds to the variable fragment of IgE. The second, called strain “D”, displays an anti-IgE DARPin, a protein able to bind to the constant part of any IgE. Allergen-specific IgE will therefore bind to both strains and clump the cells together to create an aggregate easily detectable with the naked eye or a microscope. With the serum from a blood sample of the patient, we will be able to observe aggregates if the patient is predisposed to allergies. By using a bank of allergen with as many strains A as allergens , a high-throughput screening could be performed with microfluidic strategies to identify all the allergenic predispositions of a patient. Several thousand aggregates can be sorted with these methods. This is a major break-through, which cannot be attained with current methods. Last but not least, isolated strain A can be used for desensitization, as it produces the exact allergen recognized by the patient’s IgE.