this lacuna, we have developed a flow cytometry assay to measure lympho- cyte responses to mitogens in vitro that meets clinical regulatory guidelines. (CLIA ...
S98 Abstracts
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SUNDAY
Low Values of Allergen Specific IgE Antibodies are Clinically Useful and Give Predictive Information for Very Young Children L. Soderstrom1, M. Nordlund2, C. Nilsson2, G. Lilja2; 1Phadia AB, Uppsala, SWEDEN, 2Sachs’ Children’s Hospital, Sodersjukhuset, Stockholm, SWEDEN. RATIONALE: Evaluate the diagnostic efficacy of low levels of IgE sensitisation to food and inhalant allergens among children below 5 years of age. METHODS: Two hundred and sixty-nine children, 136 boys and 133 girls, with double, maternal or no heredity of atopic disease were followed prospectively from birth to 5 years of age, for the presence of IgE sensitisation and the development of atopic manifestations. Allergen specific IgE antibody levels were measured at the age of 6, 12, 24 and 60 months to food and inhalant allergens (ImmunoCAPä, Phadia AB, Sweden), measuring range 0.1 _ 100 kU/L. RESULTS: At the age of 6 months 25% were sensitized to egg, 22% to milk, 1% to cat and no one to birch. At the age of 60 months 53% were sensitized to egg, 47% to milk, 16% to cat and 23% to birch. The low quantitative levels of specific IgE (0.1-0.35 kUA/L) were predictive for rhinitis at age 60 months of age using the birch specific IgE at age 24 months, OR 5 7.3 at 0.2 kUA/L (p 5 0.017) and predictive for eczema at 24 months using the milk specific IgE at 6 months of age, OR 5 5.6 at 0.2 kUA/L (p 5 0.007). CONCLUSION: Prospectively following levels of allergen specific IgE antibodies in children is a useful tool both as an aid in diagnosing IgE mediated allergy and for the prediction of later atopic manifestation Funding: Astma & Allergiforbundet
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Development of a Clinical Flow Cytometry Assay to Measure in Vitro Lymphocyte Responses E. C. Chenworth1, V. Nandakumar2, R. S. Abraham2; 1Division of Allergic Diseases, Mayo School of Graduate Medical Education, Rochester, MN, 2Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN. RATIONALE: T cell function is assessed in clinical laboratories using thymidine based proliferation assays. These assays evaluate bulk lymphocyte responses to mitogens and specific antigens. However, a comprehensive assessment of immune competence requires evaluation of specific subsets of cells and the number of cells entering the cell cycle. To address this lacuna, we have developed a flow cytometry assay to measure lymphocyte responses to mitogens in vitro that meets clinical regulatory guidelines (CLIA, CAP). METHODS: PBMCs from 17 healthy donors were isolated by standard protocol and run in parallel by thymidine scintillography and CFSEFACS. For each sample, a dose response curve was established to PHA, PWM and ConA in triplicate at 96 hours. RESULTS: Proliferative responses by thymidine assay were expressed as SI (stimulated raw CPM divided by unstimulated raw CPM). Similarly, for the CFSE-FACS assay, data were expressed as fold change between stimulated and unstimulated wells. A scoring system was used for comparative analysis with strong correlation ascribed to parallel dose response curves for all 4 mitogen concentrations; good correlation as 3/4, moderate correlation as 2/4 and poor correlation as 1/4. For PHA and ConA, 15/16 and 17/ 17 donors had either strong or good correlation respectively. For PWM, 9/ 16 had either strong or good correlation, and 6/16 had a moderate correlation. CONCLUSIONS: In summary, CFSE-FACS assessment of lymphocyte function is a robust alternative to thymidine assays and has the additional advantages of allowing discrimination between proliferating cell populations and cell numbers. Therefore, this assay can be expanded to include antigen specific assessment of T cell responses.
J ALLERGY CLIN IMMUNOL JANUARY 2007
386
Component Resolved IgE, IgG and IgG4 Antibody Levels for Grass Pollen Allergic Patients in Specific Immunotherapy. A Pilot Study J. Arnved1, S. Ronborg1, B. Kristensen2, L. Soderstrom3,1Lung and Allergy Clinic, Copenhagen, DENMARK, 2Sweden Diagnostics Denmark, Copenhagen, DENMARK, 3Phadia AB, Uppsala, SWEDEN. RATIONALE: To evaluate component resolved IgE, IgG and IgG4 antibody results in a group of patients undergoing grass pollen specific immunotherapy (SIT) compared to a control group of grass pollen allergic patients. METHODS: The study consisted of 36 patients getting SIT for grass pollen (active group), and 14 grass pollen allergic patients used as controls. The two groups were balanced for gender and age. Blood was drawn at the initial visit and at the follow-up occasions and tested for IgE, IgG and IgG4 antibodies for timothy (g6), and 3 components of timothy (rPhl p1, rPhl p5b, rPhl p6) using ImmunoCAPä (Phadia AB, Uppsala, Sweden). RESULTS: At the first visit no significant differences in the antibody concentrations were found, except for rPhl p1 specific IgE, where the active group had a higher antibody concentration. At visit 4 the IgG and IgG4 antibody concentration had increased for all allergens in the active group and differed significantly from those in the control group. No significant differences were found for IgE, except for rPhl p1. The proportion of IgG4 in relation to IgG increased significantly between the first visit and visit 4 in the active group, timothy 9% to 35%, rPhl p1 3% to 11%, rPhl p5b 7% to 40%, rPhl p6 3% to 11%. CONCLUSION: By using component resolved diagnostics a significant difference between the first and fourth visit in active SIT group for all allergens in the concentration of IgG and IgG4 could be demonstrated. In relation to IgG the IgG4 concentration increased more. Funding: Sweden Diagnostics Denmark
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Comparison of Skin Prick Tests and Allergen-specific IgE by MAST-CLA Technique in Allergic Thai Children P. Vilaiphan, S. Numkiatwongsa, P. Chatchatee, J. Ngamphaiboon; King Chulalongkorn Memorial Hospital, Bangkok, THAILAND. RATIONALE: The prevalence of allergic disease is increasing all over the world. In vitro IgE measurement is useful for diagnosis. Multiple allergosorbent chemiluminescent assay (MAST-CLA) is the method for detect allergen specific IgE by multipleallergen allergosorbent chemiluminescent technique using small amount of blood sample and less expensive than testing for specific IgE to each allergen separately. There have been only few studies about sensivity and specificity of MAST-CLA compared with skin prick test (SPT) in Asian population. METHODS: One hundred allergic children, aged between 3-18 years were evaluated for allergen specific IgE using MAST-CLA. The results were compared with SPT for rate of positivity, sensitivity and specificity. RESULTS: The most common aeroallergen sensitization in allergic Thai children were Dermatophagoides pteronyssinus (Dp) and Dermatophagoides farinae (Df) (88% and 87%,respectively). The most common food allergens sensitization were mixed shellfishes and shrimp (14% and 13%, respectively). The agreement between the results of MAST-CLA and SPT was best for Dp and Df with sensitivity of 85.4%. When all of the allergens in MAST-CLA panel were considered, the sensitivity rosed to 90%. The agreement was poor for food allergens. Shrimp and shellfishes, the most common food allergens in allergic Thai children were not included in MAST-CLA panel. CONCLUSIONS: MAST-CLA technique may be use to detect allergenspecific IgE. There was good correlation with SPT result especially for aeroallergens. However the panel need to be modified according to common allergens in different geographical location. Funding: Ratchadapiseksompotch Research Fund
