Polycheck

Before the COVID-19 vaccination, we determined the prevalence of SARS-CoV-2 infection using semi-quantitative anti-SARS-CoV-2 S IgG ELISA (EuroImmun GmbH, Lübeck, Germany). In addition, positive ELISA results were confirmed by quantitative anti-SARS-CoV-2 IgG immunoblots (Polycheck; Biocheck GmbH, Münster, Germany), which detected the anti-SARS-CoV-2 spike (S) protein and nucleocapsid protein (NCP) antibodies. After COVID-19 vaccination, a quantitative analysis of anti-SARS-CoV-2 S antibody levels was performed using anti-SARS-CoV-2 QuantiVac ELISA IgG (EuroImmun GmbH) according to the manufacturer’s instructions. Moreover, to analyze the SARS-CoV-2 seroprevalence after vaccination we used the anti-SARS-CoV-2 NCP IgG ELISA (EuroImmun GmbH), which detects anti-SARS-CoV-2 NCP antibodies generated only after natural infection and not produced as a consequence of BNT126b2 mRNA COVID-19 vaccination. The semiquantitative anti-SARS-CoV-2 NCP ELISA results were further confirmed by the quantitative anti-SARS-CoV-2 IgG immunoblots (Polycheck, Biocheck GmbH).

Eligible for the study were children aged 5 to 17 years with SAR diagnosed, as per current ARIA recommendations (4 (link)), with predominant sensitization to grass pollen allergens determined by skin prick testing (wheal diameter ≥ 3 mm and greater than negative control; Allergopharma-Nexter Ltd, Poland) or based on serum allergen-specific immunoglobulin E (asIgE) levels (≥ class 2; Polycheck, Biocheck GmbH, Germany), with SAR symptoms present in at least two previous grass pollen seasons. Exclusion criteria were as follows: allergy to tree pollens, receipt of PMBL within 12 months or PCBL within 6 months before the randomization visit, oral or subcutaneous allergen-specific immunotherapy within 3 years before the start of the study, treatment with systemic corticosteroids within the last 6 months before the start of the study, history of respiratory infection within 2 weeks before the randomization visit, systemic immunologic disorders, history of transfusion of blood, blood components or blood products.
Children were recruited for the study in the second half of March 2021, before the start of the grass pollen season in Poland. For all patients, written informed consent was obtained from a parent or legal guardian and assent was obtained from the patient.

The Polycheck® (Biocheck GmbH, Germany) is a multiple allergosorbent test for the quantitative measurement of allergen-specific IgE in the serum. Twenty lines of relevant allergens are coated together with five lines of calibrators on a carrier membrane, which is located in the well of the Polycheck cassette. The twenty lines represent the following respiratory allergens: pollen of plants, pets, molds, and home dust mite (HDM). Briefly, during incubation with patient's serum in the well at room temperature (RT), the allergen-specific IgE was bound to the corresponding allergens. Monoclonal ligand-labelled anti-IgE antibodies were incubated in the well cassette. Enzyme-labelled antiligand was added to the Polycheck cassettes, incubated, and bound to the immune complexes. The substrate solution was added and incubated in the dark. The color intensity of the lines was proportional to the respective allergen-specific IgE concentration in the patient's serum. The cassettes were interpreted with the Polycheck imaging software (BIS). Results were quantitatively reported in international units (kU IgE/L).