DEAMIDATED GLIADIN PEPTIDES IN COELIAC DISEASE DIAGNOSTICS

DEAMIDATED GLIADIN PEPTIDES IN COELIAC DISEASE DIAGNOSTICS

Z. Vanickova 1, P. Kocna 1, K. Topinkova 1, M. Dvorak 2 1 Institute of Clinical Biochemistry & Laboratory Diagnostics; 2 4th Medical Department, 1st Medical Faculty & General Faculty Hospital, Charles University; Prague, Czech Republic

INTRODUCTION Celiac disease (CD) is autoimmune inflammatory disease with typical histological changes of intestinal mucosa and typical antibody response. Antibodies to crude gliadin display very low specificity, which increases with purified alpha-gliadin used. Deamidated gliadin peptides (DGP) promise to increase accuracy even more. http://wheat.pw.usda.gov/ggpages/topics/gliadin.jpg http://upload.wikimedia.org/wikipedia/commons/thumb/c/ce/coeliac_path.jpg/190px-coeliac_path.jpg

INTRODUCTION Deamidated gliadin is produced by acid or enzymatic treatment of gluten. The enzyme, tissue transglutaminase, converts some of the abundant glutamines to glutamic acid. Deamidated gliadin is soluble in water. The cellular immunity to deamidated α-/β-gliadin is much greater than α/β-gliadin and can result in symptomatic gluten-sensitive enteropathy. Gliadin reactive antibodies bind very limited number of specific epitopes on the gliadin molecule. http://www.answers.com/topic/tissue-transglutaminase-png-1 http://content.answers.com/main/content/wp/en/e/e5/dqa2b5_da_gliadin.jpg

AIMS and METHODS Group of 120 serum samples from patients with small bowel biopsy performed (79 florid CD, 35 with normal histology, 6 CD in remission) was tested. We tested presence of anti-gliadin antibodies in IgA and IgG classes by: Quanta Lite Gliadin IgA II and IgG II (idgpa/g; Inova Diagnostics) Anti-gliadin ELISA IgA and IgG (edgpa/g; Euroimmun) and compared results to routinely used method: Anti-gliadin IgA and IgG (AGA/G; Dialab) All compared methods are based on ELISA principle while Inova and Euroimmun kits use DGP and Dialab kit uses purified alpha-gliadin as antigen. On the same samples also detection of immunofluorescent anti-endomysial antibodies (EmA - IgA; BioSystems) and ELISA anti-tissue transglutaminase (attg - IgA; Dialab) was performed.

Distribution of normal and positive results of routinely used methods. Histogram AGA Histogram AGG Histogram attg 0 20 40 60 80 Histogram EmA negative positive

Distribution of normal, moderate positive and positive results of methods compared for anti-gliadin antibodies in IgA class. Histogram AGA Histogram idgpa Histogram edgpa

Distribution of normal, moderate positive and positive results of methods compared for anti-gliadin antibodies in IgG class. Histogram AGG Histogram idgpg Histogram edgpg

Correlation coefficients for antigliadin antibodies in IgA class: AGA/iDGPA 0.71 AGA/eDGPA 0.19 idgpa/edgpa 0.25 idgpa 0 50 100 150 200 edgpa 0 50 100 150 200 250 300 350 edgpa 0 50 100 150 200 250 300 350 0 100 200 300 400 500 AGA 0 100 200 300 400 500 AGA 0 50 100 150 200 idgpa cut off for positivity cut off for moderate positivity

Correlation coefficients for antigliadin antibodies in IgA class: AGG/iDGPG 0.66 AGG/eDGPG 0.31 idgpg/edgpg 0.37 idgpg 0 50 100 150 200 edgpg 0 100 200 300 400 edgpg 0 100 200 300 400 0 100 200 300 400 AGG 0 100 200 300 400 AGG 0 50 100 150 200 idgpg cut off for positivity cut off for moderate positivity

sensitivity (%) specificity (%) accuracy (%) AGA 83.5 97.1 87.7 idgpa 91.1 97.1 92.3 edgpa 75.9 68.6 73.7 AGG 91.1 97.1 93.0 idgpg 87.3 100.0 91.2 edgpg 74.7 68.6 72.8 EmA 88.6 100.0 92.1 attg 86.1 97.1 89.5 Accuracy increases if IgA and IgG antibodies are evaluated together idgpa/g (96.5%), edgpa/g (78.1%). Accuracy of AGA/G combination is 94.7%.

Highest accuracies, optimal for screening by ELISA methods, are reached if anti-tissue transglutaminase and anti-gliadin antibodies are determined together. Routinely used AGA/G and attg Dialab has accuracy 95.6%, if at least one positive. Euroimmun method (edgpa/g) and attg shows only 88.6%. Inova method (idgpa/g) and attg reaches the best accuracy 96.5%.

CONCLUSION Deamidated gliadin peptides could increase sensitivities, specificities and accuracies of celiac disease serology screening. Results of this study suggest as optimal screening modality a combination of anti-tissue transglutaminase IgA and Inova idgpa and idgpg ELISA antibodies. http://www.agroweb.cz/files/image/zpravodajstvi/klasy6(dab)_450x338.jpg http://vfu-www.vfu.cz/fvhe/vegetabilie/plodiny/czech/psenice.htm