Microarrays for the Screening of Allergen-Specific IgE in Human Serum Barbara I. Fall, † Bernadette Eberlein-Ko 1 nig, ‡ Heidrun Behrendt, § Reinhard Niessner, † Johannes Ring, ‡ and Michael G. Weller* ,† Institute of Hydrochemistry, Technische Universita ¨t Mu ¨ nchen, Marchioninistrasse 17, D-81377 Mu ¨ nchen, Germany, Department of Dermatology and AllergysBiederstein, Technische Universita ¨t Mu ¨ nchen, Germany, and Division of Environmental Dermatology and Allergology GSF/TUM, Mu ¨ nchen-Neuherberg, Germany The described in vitro test system for allergy diagnosis is based on microscope glass slides activated with (3- glycidyloxypropyl)trimethoxysilane. Allergen solutions are immobilized as small droplets (∼1 0 nL) on the activated glass slides with a piezoelectric arrayer. In contrast to other tests for specific IgE, such as Pharmacia CAP FEIA, AlaSTAT, or FAST, only a 2 5 -μL serum sample is needed for the screening of allergen-specific IgE against a multi- tude of allergens and the test can be performed in less than 1 h. Compared with multiallergen dipstick screening tests (e.g., IgEquick, CMG Immunodot) based on multi- allergen-coated nitrocellulose strips, the measurement of the microarray-based system can be performed automati- cally. The chemiluminescence intensities are detected with a sensitive CCD camera. Allergen extracts and recombinant/ purified allergens (2 4 preparations) have been used on the same modified surface for the screening of allergen-specific IgE. With these disposable microarray slides, it is possible to distinguish between patients with and without elevated levels of allergen-specific IgE. Re- peated measurements of serum samples demonstrated a sufficient reproducibility. Detection limits (μg/ L) of 0 . 3 5 (r Bet v1), 0.16 (PLA 2 ), and 1.9 (Der p1) were achieved. Since the discovery of immunoglobulin E (IgE) 1,2 by Ishizaka et al. and Johansson et al. and its key role in allergic reactions, the importance of the development of test systems measuring IgE levels in serum samples has widely been recognized. In 1967, the first radioallergosorbent test (RAST) for allergen-specific IgE in serum was described. 3 Later, radioactive labels used in the RAST were replaced by many different procedures based on chromoge- nic enzyme immunoassay (EIA) or fluorescence enzyme immu- noassay (FEIA) techniques, but only a few have become routine methods for allergy diagnosis in both research and practice. 4-8 In clinical routine, the most common in vitro technique is the Pharmacia CAP System (PCS) for total and allergen-specific IgE (specific IgE FEIA, Pharmacia, Uppsala, Sweden). There are also other methods (e.g., FAST FEIA, MAST Diagnostica, Reinfeld, Germany; HYTEC EIA, Hycor Biomedicals, Kassel, Germany) commercially available. 9 Some methods are based on liquid-phase inhibitor assays (e.g., AlaSTAT, DPC Biermann, Los Angeles, CA) or multiallergen-coated nitrocellulose strips (e.g., IgEquick, Teomed AG, Greifensee, Switzerland; CMG Immunodot, Trimedal AG, Bru ¨ ttisellen, Switzerland) . 10,11 The measurements obtained with the CAP System are expressed in units (kU/ L) or in specific IgE classes (0-6). 8 The assignment of units to IgE classes 12 is shown in Table 1. Any analytical technique for the examination of specific IgE levels should be able to distinguish the classes 0 and 1. Class 0 comprises IgE concentrations of up to 0.84 μg/ L, which is equivalent to 0.35 kU/ L. The CAP System contains a cellulose polymer densely conjugated with allergen extracts or recombinant allergens. In general, allergen extracts are mixtures of different allergenic and nonallergenic proteins. 13 Carbohydrate components, e.g., from glycoproteins, are also known to be of relevance in this context. Gel electrophoresis in combination with immunoblotting techniques allowed the determination and purification of the major allergenic compounds of several allergen mixtures. 13 Recombinant DNA technology offers the possibility to produce recombinant allergenic proteins. 14 Over the past 10 years, the most important allergens from tree and grass pollens, mites, animal epithelia, insect venoms, and foods have been cloned, sequenced, and expressed. 15,16 Further studies examined the usefulness of recom- * Corresponding author. E-mail: michael.weller@ ch.tum.de. † Institute of Hydrochemistry, Technische Universita ¨tMu ¨ nchen. ‡ Department of Dermatology and AllergysBiederstein, Technische Univer- sita ¨tMu ¨ nchen. § Division of Environmental Dermatology and Allergology GSF/ TUM. 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