Recombinant allergens and their application in the diagnostics of allergies

LA_Apronex-1.jpgAllergies since first description of pollen allergy “hay fever” in 1870 manifest as a system diseases triggered by our immune system hyperactivity to various foreign substances, in which epithelial barrier plays an important role in allergy sensitization and tolerance. Allergy as a chronic disease is largely accompanied by IgE-mediated immune response, activation and degranulation of basophils and inflammation, in most severe outcomes leading to life-threatening symptoms/disease such as anaphylactic shock or asthma – reviewed in (1,2).

Taking in account both life-complicating (atopic dermatitis and rhinitis) and life-threatening manifestations of allergy a significant effort was especially in last 30 years invested into both relevant diagnosis and effective treatment of mainly though not exclusively life-endangering allergies (not just their symptoms). The most common and widely used allergy diagnostic tests are based on the skin allergy response and include skin prick, intradermal and patch tests mainly still using complex allergens/extracts (3). In vitro diagnostic approaches (ELISA, multiplex CHIP assays such as Alex or Immunocap ISAC) are based on the detection and quantification of the levels of relevant allergens-specific IgE antibodies and employ multiple spotted complex and especially recombinant allergens (4). Their disadvantages might be however high cost and likely seasonal and time-related variations in the levels of distinct allergens-specific IgE antibodies. In addition to these two major diagnostic approaches there is a semi-in vitro diagnostic assay using both complex and preferably recombinant allergens, which is based on the activation of basophils – Basophil Activation Test (BAT) (5).

BAT.jpgPrinciple of BAT: After adding various allergens to the peripheral blood samples of particular patient, the basophils (identified by a specific marker, in this case CD203c in PE channel) react by degranulation only to the causal allergen. Degranulation can be detected by flow cytometry as externalization of CD63 marker (in this case in FITC channel), and easily distinguished from resting basophils.

Both multiplex CHIP assays as well as BAT tests exploit recombinant protein allergens as reliable and reproducible reagents (in contrast to natural allergens with batch-to-batch variations and higher complexity) (6). Since the preparation of the first recombinant allergen Der p 1 from dust mite in 1988 (7), their number skyrocketed and currently there is known over 500 recombinant allergens listed in several databases such as WHO/IUIS Allergen , Allergome and other (8). Recombinant allergens are produced both in prokaryotic (mainly E. coli) and in eukaryotic (yeast and insect cells) expression systems and in addition to their use in diagnostics find their way also into immunotherapy of various allergies (9). For example ImmunoCAP ISAC multiplex assay contains 112 various recombinant allergens and ALEX multiplex assay 125 recombinant allergens (10). The recombinant allergens (single or relevant combinations of allergens) can be in addition to IgE levels quantifying multiplex assays also exploited in BAT assays (11) as the recombinant honey bee rApi m 1, wasp rVes v 5 were used in BAT assay for distinguishing false double positivity from IgE assays (12). Exbio together with BAT assay kit BasoFlowEx offers a growing list of the recombinant allergens produced by a sister company Apronex and allergen extracts - see list below.

Ladislav Andera
Institute of Biotechnology AS CR and Apronex s.r.o.

BasoFlowEx kit
T-cell BlastoFlowEx kit
Allergens intended for Basophil Activation Test (BAT) 

Recombinant allergens (ELISA, FC application)
Sensitivity tests


1. Dougherty, J. M., Alsayouri, K., and Sadowski, A. (2020) Allergy. in StatPearls, Treasure Island (FL). pp
2. Krempski, J. W., Dant, C., and Nadeau, K. C. (2020) Ann Allergy Asthma Immunol 125, 507-516
3. Birch, K., and Pearson-Shaver, A. L. (2020) Allergy Testing. in StatPearls, Treasure Island (FL). pp
4. Buzzulini, F., Da Re, M., Scala, E., Martelli, P., Conte, M., Brusca, I., and Villalta, D. (2019) Clin Chim Acta 493, 73-78
5. Hemmings, O., Kwok, M., McKendry, R., and Santos, A. F. (2018) Curr Allergy Asthma Rep 18, 77
6. Valenta, R., Karaulov, A., Niederberger, V., Zhernov, Y., Elisyutina, O., Campana, R., Focke-Tejkl, M., Curin, M., Namazova-Baranova, L., Wang, J. Y., Pawankar, R., and Khaitov, M. (2018) J Allergy Clin Immunol Pract 6, 1845-1855 e1842
7. Thomas, W. R., Stewart, G. A., Simpson, R. J., Chua, K. Y., Plozza, T. M., Dilworth, R. J., Nisbet, A., and Turner, K. J. (1988) Int Arch Allergy Appl Immunol 85, 127-129
8. Radauer, C., and Breiteneder, H. (2019) Allergy 74, 2057-2060
9. Tscheppe, A., and Breiteneder, H. (2017) Int Arch Allergy Immunol 172, 187-202
10. Melioli, G., Puggioni, F., Racca, F., Descalzi, D., Canonica, G. W., and Heffler, E. (2019) Curr Opin Allergy Clin Immunol 19, 204-208
11. Eberlein, B. (2020) Front Immunol 11, 1815
12. Eberlein, B., Krischan, L., Darsow, U., Ollert, M., and Ring, J. (2012) J Allergy Clin Immunol 130, 155-161