Eur J Pediatr (1990) 150: 37-39 European Journal of Pediatrics 9 Springer-Verlag1990 Screening for delayed-hypersensitivity in Italian children: multicentric study by multitest skin testing M. Bardare 1, L. Armenio 2, L. Businco 3, G. Cavagni 4, M. De Martino s , M. Duse 6, M. Masi 7, V. Monafo s, M. C. Pietrogrande 1, P. Tovo 9, and M. C. Venturi 1 1First Paediatric Department, University of Milano, Via Commenda 9, 1-20122 Milano, Italy 2Second Paediatric Department, University of Bari, Italy 3Third Paediatric Department, University of Rome, Italy 4paediatric Department, University of Parma, Italy 5 Second Paediatric Department, University of Firenze, Italy 6paediatric Department, University of Brescia, Italy 7Third Paediatric Department, University of Bologna, Italy SPaediatric Department, University of Pavia, Italy 9First Paediatric Department, University of Torino, Italy Received August 31, 1989 / Accepted February 6, 1990 Abstract. The Multitest skin test allows the simultaneous intradermal application of seven standardized recall anti- gens in a reproducible manner. Response reading is based on a scoring system that distinguishes between normal and diminished delayed hypersensitivity (DH). To de- termine incidence and size of DH responses for each an- tigen in relation to age and sex in healthy Italian children, a multicentric study was performed on a representative paediatric population of 491 subjects (285 males and 296 females) aged from 4 months to 16 years. In both sexes DH reactivity as measured by score and number of posi- tive responses increased significantly with age. The inci- dence of anergy was 8.8% for the whole population and decreased significantly with age in both sexes. Incidence of positive responses was highest for tetanus and diphteria toxoids, intermediate for Candida, Proteus and Strepto- coccus, and lowest for tuberculin and Trichophyton anti- gens. There was no significant difference between sexes in the incidence of positive response to each antigen. Key words: Delayed hypersensitivity - Skin testing - Cell-mediated immunity Introduction In the assessment of cell-mediated immunity (CMI) in vivo tests are very useful because they give an overall representation of the total CMI response to antigens. Until recently delayed hypersensitivity (DH) skin tests have suffered from the small number of antigens used, the lack of standardisation of testing methods and the Offprint requests to: M. Bardare Abbreviations: CMI = cell-mediated immunity; DH = delayed hypersensitivity subsequent incorrect interpretation of results [1, 3, 5, 9]. A multiple skin test has recently been developed to per- mit rapid simultaneous intradermal application of seven standardized recall antigens. The reading of responses is based on a scoring system, useful in distinguishing be- tween normal DH reactivity and decreased response. To provide normal values in relation to age and sex, many studies on healthy adults were undertaken in the United States [4] and in Europe [3, 6-8] but only one study was performed in schoolchildren in the United States [2]. The aim of the present study is to provide normal val- ues for DH reactivity in a healthy Italian paediatric pop- ulation. Materials and methods Population studied The population studied was composed of 491 healthy children, 285 males and 296 females, aged between 4 months and 16 years (aver- age age 62 months), from nine different paediatric centres in Italy: Milan, Turin, Brescia, Pavia, Parma, Bologna, Florence, Rome, Bari. For each child tested, age, sex, diphtheria, tetanus and tuber- culin immunisations were recorded. The children were divided into three age groups: 4 months - 2 years (group 1) 2-6 years (group 2), 6-16 years (group 3). Group 1 was composed of 159 subjects, 87 males and 72 females (median age 17.3 months); group 2 was composed of 175 subjects, 105 males and 70 females (median age 51.7 months), and group 3 contained 157 subjects, 93 males and 64 females (median agze118.4 months). Sex distribution was similar in all three groups (Z = 2.04, P < 0.05). Testing procedure and measurement of response The Multitest CMI (Instituto Merieu• Rome) was used. This is a methylpolymetacrylate applicator with eight heads each with nine tines loaded with sterile antigen solution. The following seven anti- gens, obtained from inactivated culture supernatants and stabilized with a 70% w/v glycerine solution, were used: toxoid from Clos-