Assessment of the Sensitizing Potential of Proteins in BALB/c Mice: Comparison of Three Protocols of Intraperitoneal Sensitization

Abstract

Most food allergy cases are associated with a limited group of allergens. This could be attributed to an increased ability of some foods to sensitize and trigger allergic reactions. However, there are no validated animal models to evaluate the sensitizing or allergenic potentials of proteins. Our aim was to evaluate three protocols of adjuvant-free intraperitoneal sensitization that differ in the time points for sample collection (days 14, 28 and 35 from beginning of the sensitization) and also in the number of immunizations (2, 5 and 3, respectively). Ovalbumin (OVA; 0.05 mg), cow milk proteins (CMP; 0.025, 0.05 and 0.25 mg), and potato acid phosphatase (PAP; low allergenic protein; 250.0 mg) were administered intraperitoneally (ip) to BALB/c mice (n = 4–6) and the protein-specific IgE and IgG antibody responses were evaluated using ELISA. Additional serum protein-specific IgE antibodies evaluations were carried out after IgG depletion. Anti-OVA IgE antibodies were detected in mice from all three protocols. The responses were higher in the group of mice that underwent the 28-day protocol than in those that underwent the 14- or 35-day protocols (p < 0.01 and p < 0.05, respectively). Anti-CMP IgE antibodies were detected in both the 14- and 28-day protocols, but the response was higher in the group that underwent the 28-day protocol (p < 0.001). The anti-CMP IgE antibody response detection was improved after serum IgG depletion (p < 0.001). Anti-PAP IgE antibodies were not detected. Mice with undetectable serum levels of protein-specific IgE triggered anti-OVA, -CMP, and -PAP IgG responses. An adjuvant-free 28-day protocol with five ip immunizations seems appropriate for evaluation of the inherent sensitizing or allergenic capacity of the studied proteins. Reproducible results were obtained utilizing the BALB/c mouse strain. Inter-laboratory studies including a larger number of proteins should be carried out to validate this model.