**2. Diagnostic tools and monitoring**

and Allergies Phase Three. The study included around 14,000 from 5 regional centers different geographical and urban characteristics (children both from urban and rural areas participated). Investigators analyzed the prevalence of allergic diseases in two age groups (the first one preschool children aged 6–7 years old and the second one children between the age of 13–14 years old. The prevalence of asthma was 6.59% in younger age group, whereas the prevalence in older age group was around 5.36%. Note that 7.17% of preschool children and 14.89% of school children were diagnosed allergic rhinitis. Overall, asthma prevalence was 5.91%, rhinitis 11.46%, and eczema 14.27% [2]. The growing worldwide burden of allergic diseases is properly defined as the "allergy epidemic." The German epidemiological Multicenter Allergy Study (MAS) suggested an age-related evolution of atopic and allergic diseases, usually named "atopic march." In fact, on epidemiological bases, infantile eczema and food allergy usually precede the onset of allergic airway disease (rhinitis and asthma). It is also interesting to point out that unlike other common chronic diseases such as diabetes mellitus or hypertension, it is well established that the development of allergic diseases start just after birth or according to some authors maybe earlier in prenatal period [3]. The incidence of asthma is the highest in preschool and early school age with an improvement in symptoms and a decrease in prevalence afterwards, but with one more pic in incidence in adolescents' period especially in female teenagers mainly due to hormone disturbance. It is well known that allergic diseases are multi factorial which means that in their pathophysiology both genetic and environmental factors are included. Atopic family history is one of the most important risk factors for the development of asthma. MAS cohort study analyzed the main risk factors for persistent asthma/wheeze in an early adolescent's period. According to the results from this huge study wheezing before the age of 3 as well as wheezing after the age of 6, accompanied with early atopic dermatitis, positive family history of atopic diseases and positive allergy tests, particular to perennial allergens represent the main

Although according to birth cohort studies data we are aware that genetic burden has an important influence in allergies development and despite lots of efforts, we have still failed to identify responsible genes. Many factors in the environment contribute to the development of allergies (e.g., diet, immunizations, antibiotics, pets, and tobacco smoke), but we do not know how to modify the environment to reduce the risks [6]. According to several epidemiological studies, a decline in microbial diversity was proposed to have an important role in allergic epidemic, best summarized in hygiene hypothesis, and nowadays defined as "biodiversity hypothesis." Identification of prenatal and early postnatal risk factors is of a great importance for early prevention and successful intervention. Two recent studies showed that reduce food diversity in early childhood can be associated with atopic sensitization andallergic diseases later on. It is also suggested that high "antigen burden" in early life can be a protective factor necessary to "educate" the immune system and to prevent childhood allergic diseases. Early allergy prevention that includes: administrations of probiotics to pregnant mothers and to high-risk children, oral or intranasal extracts, and earlier introduction of foods is still matter of a debate due to conflicting results [7, 8]. Despite many different options are currently available for the diagnostic workup and management, the burden of allergic airway

risk factors [4, 5].

102 Allergen

Despite many different diagnostic tools for allergic disease it still remains a challenge especially in infants and toddlers. Skin tests represent an important diagnostic tool in workup of many allergic diseases. These tests are mainly used for the diagnosis of inhalant allergies, but nowadays there are more and more tendencies to use this kind of tests for allergies to food, venom, occupational agents, and drugs. Skin prick tests (SPTs) and intradermal tests still represent the cornerstone of the diagnosis of IgE-mediated (type I) allergies. They are easy to perform, cheap and allow a fast reading, usually performed in outpatient clinics. Performing skin prick tests needs a specific training, especially for intradermal and epicutaneous tests with nonstandard allergens, that are not usually performed in children population. Special precautions that have to be considered before performing skin prick tests include the usage of some drugs, skin conditions and in adolescents and adults pregnancy. Before performing *in vivo* skin prick tests patients are not allowed to take drugs such as antihistamines at least several days because it is well known that these kinds of drugs could mask positive results of type I reactions, on the other side conditions like pressure urticaria or dermographism are able to provoke false positive results. For that reason using positive control histamine and negative control saline solution are crucial for results interpretations. Skin prick tests (SPT) are one of the most important diagnostic tools in asthma and AR diagnosis with sensitization to inhalant allergens. They can get prompt information on sensitization to inhalant allergens such as pollen, house dust mites, pets, to a lesser extent molds. Recommendation for SPTs is available with more or less variation in many climate and geographical areas. As they are very cheap and easy to perform SPTs are of a great importance especially in undeveloped or developing countries. Here, it is also interesting to mention that in tropical areas standard SPTs battery should include typical tropical allergens such as Blomia tropical. In southeastern and Western Europe standard allergens for preforming SPTs usually include following allergen solutions: tree, ragweed, and grass pollen, house dust mite, molds, cockroach, dogs, and cats dander. Before starting allergen-specific immunotherapy SPTs have to be performed [10–14]. Nasal and bronchial provocation test are indicated for patients with typical clinical symptoms and signs of allergic rhinitis and/ or asthma but with negative *in vivo* skin prick tests [15]. Those tests should be performed exclusively by a well-trained staff at the allergy departments. They are very important for distinguishing allergic and nonallergic rhinitis as well as for the diagnosis of local allergic rhinitis (LAR)—typical clinical history of allergic rhinitis with positive nasal provocation test, usually with elevated eosinophils in nasal smear, but with negative skin prick or/and *in vitro* allergy tests [16]. *In vitro* allergy tests are cornerstone of allergy diagnostic especially in the pediatric population. All children with positive clinical history of allergic diseases (atopic dermatitis, allergic rhinitis, and/or asthma) should be evaluated, particularly those with positive uni- or bilateral family history of atopic diseases. Determinations of total IgE, followed with evaluation of the allergen-specific antibody levels are precede the introduction of allergen-specific immunotherapy. It is also a very important to be mentioned that the interpretation of the allergy tests should be strictly done in the light of clinical history of a certain patients. Novel diagnostic tools are also capable to determine sensitization to a specific pure or recombinant allergens that is of a great importance for individualized treatment approach. Sometimes this kind of tests are the most relevant to confirm the diagnosis of allergy sensitization. To data, the most commonly used system to determine allergen-specific IgE is the ImmunoCAP system that are considered as a goal standard for *in vitro* diagnosis of allergen condition. Despite great technological improvements in *in vitro* diagnostics of allergies, several problems still remain. Although elevated IgE is a marker of IgE-mediated allergy, this is not sufficient for the induction of symptoms. According to the data, more than 20% of patients with elevated IgE are in fact asymptomatic. Elevated serum IgE level is irrelevant as long as it does not bind to Fcέ receptors on effectors cells (mastocytes, eosinophils, and basophils). Positive allergen-specific IgE in serum is not sufficient to confirm allergy in all cases [17–19]. At the current state of art it is a very important to be a little bit septic about allergy diagnostic test results only based on determination of allergen-specific serum IgE levels and to consider clinical history, accompanied by adequate skin prick tests or provocation tests, which drive the diagnosis before considering allergen-specific immunotherapy inclusion. During the last decades, there has been a huge improvement in *in vitro* allergy diagnosis due to novel approaches that include molecular components. It has been already mentioned that allergen-specific tests are not enough sensitive and specific for allergy diagnosis, through the advent of molecular technology, some weaknesses, and shortcomings of classical approach that used only natural extracts could be solved. Component resolved diagnosis (CRD) of the specific IgE response provides more individual approach in diagnosis of allergic patients and better selection of patients for allergen-specific immunotherapy. It is also of a great value for monitoring of the efficacy, immunogenicity and safety of allergenspecific immunotherapy [20, 21]. Cellular allergy testing represents one more *in vitro* allergy diagnostic tool. It expands the tools of allergist to diagnose and monitor allergic diseases. The basophil activation test (BAT) is the most common used cellular allergy tests in routine clinical practice and in research. That test is able to document type I sensitization to a specific allergen, as fraction of blood basophils activated by soluble allergen. Basophil sensitivity can be used for identification the main sensitizer among cross-reacting allergens or allergen preparation as well as for monitoring progress of allergen-specific immunotherapy and anti-IgE therapy [22–24].
