Ranking of immunodominant epitopes in celiac disease: Identification of reliable parameters for the safety assessment of innovative food proteins.

A ranking of gluten T-cell epitopes triggering celiac disease (CD) for its potential application in the safety assessment of innovative food proteins is developed. This ranking takes into account clinical relevance and information derived from key steps involved in the CD pathogenic pathway: enzymatic digestion, epitope binding to HLA-DQ receptors of the antigen-presenting cells and activation of pro-inflammatory CD4 T-cells, which recognizes the HLA-DQ-epitope complex and initiates the inflammatory response. In silico chymotrypsin digestion was the most discriminatory tool for the ranking of gluten T-cell epitopes among all digestive enzymes studied, classifying 81% and 60% of epitopes binding HLA-DQ2.5 and HLA-DQ8 molecules, respectively, with a high risk. A positive relationship between the number of prolines and the risk of gluten T-cell epitopes was identified. HLA-binding data analysis revealed the additional role played by the flanking regions of the 9-mer epitopes whereas the integration of T-cell activation data into the ranking strategy was incomplete because it was difficult to combine results from different studies. The overall ranking proposed in decreasing order of immunological relevance was: α-gliadins > ω-gliadins > hordeins > γ-gliadins âˆ¼ avenins âˆ¼ secalins > glutenins. This novel approach can be considered as a first step to reshape the risk assessment strategy of innovative proteins and their potential to trigger CD.

Is Endoscopic Assessment of the Esophagus and Stomach Enough to Determine the Need for Biopsy at These Sites in Pediatric Patients Undergoing Endoscopy for Elevated TTG?

OBJECTIVES: The goal of our study was to determine whether visual assessment of the esophagus and stomach could predict abnormal histology and determine the frequency of interventions based on biopsies in patients undergoing endoscopy for elevated tissue transglutaminase immunoglobulin A antibody (TTG). METHODS: Pathology records were searched for patients with biopsy performed for elevated TTG. Pathology report, endoscopy report, and follow-up were obtained and slides from the duodenum reviewed. Pathology was considered gold standard for sensitivity and specificity calculations. RESULTS: 240 patients were included. 215 patients had esophageal biopsies performed. Esophageal endoscopic visual assessment had sensitivity of 47% and specificity of 93% for abnormal histology. 16(7%) patients had therapy or referral related to results and, of these, 6(38%) had visually normal endoscopy. 237 biopsies were performed of stomach. Gastric endoscopic visual assessment had a sensitivity and specificity of 20% and 87%. 24(10%) patients had therapy based on findings and, of these, 12 (50%) had visually normal endoscopy. CONCLUSIONS: Endoscopic assessment of esophagus and stomach has low sensitivity and high specificity for pathologic abnormalities when indication for endoscopy is elevated TTG. When endoscopy is visually normal clinical interventions based on biopsy are rare, and foregoing biopsy may be considered.

Assessment of European Society of Paediatric Gastroenterology Hepatology and Nutrition (ESPGHAN) guidelines in an Australian paediatric population.

Coeliac disease (CD) diagnosis is based on clinical assessment, detection of specific autoantibodies and histological examination of small intestinal biopsies. The European Society of Paediatric Gastroenterology Hepatology and Nutrition (ESPGHAN) guidelines have recently been updated and recommend CD may be diagnosed without a biopsy or HLA typing in symptomatic patients with high titre IgA tissue transglutaminase antibodies (aTTG) and positive endomysial antibodies (EMA). However, the need for EMA in patients with high level aTTG has been questioned. We aimed to determine the diagnostic benefit of HLA typing, EMA and IgG antibodies to deamidated gliadin (DGP) in children with high level aTTG. We prospectively evaluated children presenting for assessment of possible CD. All patients underwent small bowel biopsy, serological testing and HLA typing. Results were analysed and correlated with histopathological diagnosis. A total of 209 children were assessed; 61.5% were found to have CD and 29% could have avoided biopsy as per 2020 ESPGHAN guidelines. Titres of aTTG ≥60 U/mL or DGP ≥28 U/mL gave 100% specificity and 100% positive predictive value (PPV) for CD. HLA typing and EMA did not improve the PPV of patients with aTTG ≥60 U/mL, but addition of DGP ≥28 U/mL improved diagnostic sensitivity whilst retaining 100% specificity. Addition of HLA and EMA testing in patients with high titre aTTG antibodies does not improve diagnostic performance and may possibly be omitted from the serological workup in these patients. Our data support combining aTTG and DGP testing and optimising cut-offs to maximise specificity as an alternative biopsy-free diagnostic approach.

An integrated, accurate, rapid, and economical handheld consumer gluten detector.

Celiac disease, characterized by autoimmune reactions to dietary gluten, affects up to 3 million in the US and approximately 0.5%-1% globally. A strict, lifelong gluten-free diet is the only treatment. An economic, simple, accurate, rapid and portable gluten testing device would enable gluten-sensitive individuals to safeguard their food safety. We developed a novel solution, Nima™, a gluten sensor that integrates food processing, gluten detection, result interpretation and data transmission in a portable device, detecting gluten proteins at or below the accepted 20 ppm threshold. We developed specific monoclonal antibodies, an optimized lateral flow immunoassay strip, and one-step aqueous extraction. Compared with reference R5, NimaTM antibodies (13F6 and 14G11) had 35- and 6.6-fold higher gliadin affinities, respectively. We demonstrated device performance using a comprehensive list of foods, assessing detection sensitivity, reproducibility, and cross-reactivity. Nima™ presented a 99.0% true positive rate, with a 95% confidence interval of 97.8%-100%.

Diagnostic Yield of 2 Strategies for Adult Celiac Disease Identification in Primary Care.

GOALS: To compare the diagnostic yield and cost-consequences of 2 strategies, screening regardless of symptoms versus case finding (CF), using a point-of-care test (POCT), for the detection of celiac disease (CD) in primary care, to bridge the diagnostic gap of CD in adults. MATERIALS AND METHODS: All subjects under 75 years of age who consecutively went to their general practitioners' offices were offered POCT for anti-transglutaminase immunoglobulin A antibodies. The POCT was performed on all subjects who agreed, and then a systematic search for symptoms or conditions associated with higher risk for CD was performed, immediately after the test but before knowing the test results. The 2 resulting groups were: (a) POCT positive and (b) symptomatic subject at CF. Subjects were defined as symptomatic at CF in the presence of 1 or more symptoms. All POCT-positive or symptomatic subjects at CF were referred to the CD Centers for confirmation of CD. Data on resource consumption were gathered from patients' charts. Cost of examinations, and diagnostic and laboratory tests were estimated with regional outpatient tariffs (Sicily), and a price of &OV0556;2.5 was used for each POCT. RESULTS: Of a total of 2197 subjects who agreed to participate in the study, 36 (1.6%) and 671 (30.5%) were POCT positive and symptomatic at CF, respectively. The yield from the screening and CF was 5 new celiac patients. The total cost and mean cost for each new CD case were &OV0556;7497.35 and &OV0556;1499.47 for the POCT screening strategy, and &OV0556;9855.14 and &OV0556;1971.03 for the CF strategy, respectively. Assuming consecutive use of both strategies, performing POCT only in symptomatic subjects at CF, the calculated yield would be 4 new diagnoses with a total cost of &OV0556;2345.84 and a mean cost of &OV0556;586.46 for each newly diagnosed patient. Only 1 patient was celiac despite a negative POCT. CONCLUSIONS: Testing symptomatic subjects at CF only by POCT seems the most cost-effective strategy to bridge the diagnostic gap of adult CD in primary care.

Allergenicity Assessment of Transgenic Wheat Lines In Silico.

Agriculture biotechnology is a promising tool for developing varieties with enhanced quality and quantity. Transgenic proteins expressed by genetically modified (GM) food crops improve crop characteristics like nutritional value, taste, and texture, and endow plants with resistance against fungus, pests, and insects. Despite such potential benefits, there are concerns regarding possible adverse effects of GM crops on human health, animals and the environment. Among the proposed guidelines for GM food safety testing-the weight-of-evidence approach proposed by the Codex Alimentarius Commission (ALINORM 03/34A) is the most recent. Till date, several transgenic wheat lines have been developed and research is underway for further improvement. However, GM wheat is not being grown or consumed in any part of the world. In the present study, in silico tools were employed for safety testing of eight transgenes used for the development of transgenic wheat lines. Among the genes studied, none of them shared sequence homology with the reported allergens and may be safe for use in genetic engineering. In conclusion, gene selection for developing transgenic wheat lines should be done with utmost care to ensure its safety for feed and fodder.

Strategies to improve the efficiency of celiac disease diagnosis in the laboratory.

The demand for testing to detect celiac disease (CD) autoantibodies has increased, together with the cost per case diagnosed, resulting in the adoption of measures to restrict laboratory testing. We designed this study to determine whether opportunistic screening to detect CD-associated autoantibodies had advantages compared to efforts to restrict testing, and to identify the most cost-effective diagnostic strategy. We compared a group of 1678 patients in which autoantibody testing was restricted to cases in which the test referral was considered appropriate (G1) to a group of 2140 patients in which test referrals were not reviewed or restricted (G2). Two algorithms A (quantifying IgA and Tissue transglutaminase IgA [TG-IgA] in all patients), and B (quantifying only TG-IgA in all patients) were used in each group, and the cost-effectiveness of each strategy was calculated. TG-IgA autoantibodies were positive in 62 G1 patients and 69 G2 patients. Among those positive for tissue transglutaminase IgA and endomysial IgA autoantibodies, the proportion of patients with de novo autoantibodies was lower (p=0.028) in G1 (11/62) than in G2 (24/69). Algorithm B required fewer determinations than algorithm A in both G1 (2310 vs 3493; p<0.001) and G2 (2196 vs 4435; p<0.001). With algorithm B the proportion of patients in whom IgA was tested was lower (p<0.001) in G2 (29/2140) than in G1 (617/1678). The lowest cost per case diagnosed (4.63 euros/patient) was found with algorithm B in G2. We conclude that opportunistic screening has advantages compared to efforts in the laboratory to restrict CD diagnostic testing. The most cost-effective strategy was based on the use of an appropriate algorithm.

Implementation of National Institute for Health and Care Excellence (NICE) guidance to measure immunoglobulin A with all coeliac screens: can an affordable solution be devised?

There has been a dramatic increase in requests for coeliac disease (CD) serological screening using immunoglobulin (Ig)A tissue transglutaminase antibodies (IgA-tTG). Recently, the UK National Institute for Health and Care Excellence has revised its guidance, recommending that total IgA should also be measured in all samples. This is justified, as false-negative results may occur with IgA deficiency. However, implementation of this guidance will incur considerable expense. Tests that measure IgA-tTG antibodies can detect IgA deficiency, indicated by low background signal. This provides an opportunity to identify samples containing IgA ≤ 0·2g/l, obviating the need for unselected IgA measurement. We investigated the feasibility of this approach in two centres that use the EliA™ Celikey™ assay or QUANTA Lite® enzyme-linked immunosorbent assay to quantify IgA-tTG antibodies. In both cases, total IgA correlated strongly with background IgA-tTG assay signal. Using the Celikey™ assay, a threshold of < 17·5 response units achieved 100% sensitivity (95% confidence intervals 79·4-100%) for detection of IgA ≤ 0·2g/l, circumventing the need for IgA testing in > 99% of sera. A similar principle was demonstrated for the QUANTA Lite® assay, whereby a threshold optical density of < 0·0265 also achieved 100% sensitivity (95% confidence intervals 78·2-100%) for IgA ≤ 0·2 g/l, avoiding unnecessary IgA testing in 67% of cases. These data suggest that CD screening tests can identify samples reliably containing low IgA in a real-life setting, obviating the need for blanket testing. However, this approach requires careful individualized validation, given the divergent efficiency with which assays identify samples containing low IgA.

HLA genotyping as first-line screening tool for coeliac disease in children with juvenile idiopathic arthritis.

OBJECTIVES: Coeliac disease (CD) and juvenile idiopathic arthritis (JIA) often coexist. This association warrants assessment for CD in patients with JIA. We evaluated the clinical relevance and cost-effectiveness of human leucocyte antigen (HLA) genotyping in first-line screening for development of CD in children with JIA. PATIENTS AND INTERVENTIONS: 95 patients with JIA were screened for CD using CD-specific antibodies. In case of positivity, a small intestinal biopsy was performed to confirm diagnosis. In addition, HLA genotyping was performed. 110 age-matched and sex-matched Caucasian children from the same geographical area served as controls. RESULTS: CD was diagnosed in 4 of 95 patients with JIA (4.2%), a rate significantly higher compared with controls (p<0.02) and 14 times higher than in the general population. Twenty-six patients (27.4%) had one of the variants of the risk genotypes. All four patients diagnosed with CD had a HLA-DQ2.5 genotype: one was homozygote, the remainder heterozygote. Twenty-two patients are, judging by their HLA genotypes, at risk of developing CD and require repeated serological screening. None of the 69 patients without HLA-DQ2/DQ8 genotypes had CD-specific antibodies. Screening with HLA genotyping becomes cheaper than screening without after the second determination. CONCLUSIONS: In our cohort of patients with JIA, lack of HLA-DQ2/DQ8 genotypes identified a majority not at risk of CD in whom repeated serological testing is unnecessary. Genotyping is nowadays the most efficient and cost-effective way to screen for CD risk in JIA.

The clinical and phenotypical assessment of seronegative villous atrophy; a prospective UK centre experience evaluating 200 adult cases over a 15-year period (2000-2015).

BACKGROUND: Seronegative villous atrophy (SNVA) is commonly attributed to coeliac disease (CD). However, there are other causes of SNVA. More recently angiotensin-2-receptor-blockers (A2RBs) have been reported as an association but data on SNVA have been limited to centres evaluating complex case referrals and not SNVA in general. OBJECTIVES: To provide clinical outcomes and associations in a large prospective study overseeing all newcomers with SNVA. DESIGN: Over a 15-year period (2000-2015) we evaluated 200 adult patients with SNVA at a UK centre. A diagnosis of either seronegative CD (SNCD) or seronegative non-CD (SN-non-CD) was reached. Baseline comparisons were made between the groups, with 343 seropositive CD subjects serving as controls. RESULTS: Of the 200 SNVA cases, SNCD represented 31% (n=62) and SN-non-CD 69% (n=138). The human leucocyte antigen (HLA)-DQ2 and/or DQ8 genotype was present in 61%, with a 51% positive predictive value for SNCD. The breakdown of identifiable causes in the SN-non-CD group comprised infections (27%, n=54), inflammatory/immune-mediated disorders (17.5%, n=35) and drugs (6.5%, n=13; two cases related to A2RBs). However, no cause was found in 18% (n=36) and of these 72% (n=26/36) spontaneously normalised duodenal histology while consuming a gluten-enriched diet. Following multivariable logistic regression analysis an independent factor associated with SN-non-CD was non-white ethnicity (OR 10.8, 95% CI 2.2 to 52.8); in fact, 66% of non-whites had GI infections. On immunohistochemistry all groups stained positive for CD8-T-cytotoxic intraepithelial lymphocytes. However, additional CD4-T helper intraepithelial lymphocytes were occasionally seen in SN-non-CD mimicking the changes associated with refractory CD. CONCLUSIONS: Most patients with SNVA do not have CD, in particular those who are not white. Furthermore, a subgroup with no obvious aetiology will show spontaneous histological resolution while consuming gluten. These findings suggest caution in empirically prescribing a gluten-free diet without investigation.

Challenges in the Standardization of Autoantibody Testing: a Comprehensive Review.

Standardization and harmonization are complementary tools to achieve higher testing quality in laboratory medicine. Both are of great relevance and are strongly needed in autoimmune diagnostics, due to the impressive advance in basic research and technological development observed in this diagnostic field in recent years that has led to the introduction of many new tests and new analytical methods. It is, therefore, essential that this strong innovative thrust is translated into clinical practice in a coordinated way to avoid confusion and the risk of potentially harmful errors for the patient. However, while standardization of antibody assays is a very complex task, harmonization of procedures and behaviors is a more feasible target and should necessarily include all the phases of the total testing process-in the pre-analytical phase, appropriateness of test requests, harmonization of autoantibody terminology, and adoption of uniform nomenclature for laboratory tests; in the analytical phase, harmonization of measurements, and sharing of test profiles and diagnostic algorithms; and in the post-analytical phase, harmonization of data reporting, and criteria for interpreting immunoserological results, especially harmonization of units, reference intervals, decision limits, and definition and notification of critical values. We here provide and discuss some examples of harmonization initiatives related to anti-nuclear antibodies, TSH receptor, and anti-thyroid peroxidase antibodies and to antibodies associated with autoimmune hepatitis and with celiac disease. These initiatives could be the starting steps to achieve a wider consensus and a closer interaction among stakeholders in the path of autoimmune diagnostics harmonization to enhance clinical effectiveness and provide greater patient safety.

Oat raw materials and bakery products - amino acid composition and celiac immunoreactivity.

BACKGROUND: The aim of this study was to compare the biochemical and immunochemical properties of avenins in some special oat raw materials and additionally the possibility of using them as a raw material for the gluten-free bakery products. METHODS: The compared oat raw materials were - oat flakes, commercial oat flours (including gluten-free oat flour) and residual oat flour, which is by-product of ß-glucan preparation. Biochemical characteristic included amino acid compositions and SDS-PAGE profiles of extracted avenins. The immunochemical reactivity with polyclonal anti-gluten and monoclonal anti-gliadin antibodies was evaluated qualitatively and quantitatively by immunoblotting and ELISA methods. Additionally, experimental bakery products made of examined raw materials were assessed according to their suitability for the celiac patients' diet. RESULTS: The highest protein content was measured in the ß-glucan preparation "Betaven" and gluten-free oat flour. Proteins of all materials are rich in glutamic and aspartic acid, leucine and arginine. Proportions of amino acids in avenins extracted from most of oat raw materials are similar, excluding gluten-free oat flour, which has a very low avenin content and proportions of individual amino acids are different. The SDS-PAGE protein pattern consisted of proteins with molecular weight of about 25-35 kDa. Polyclonal anti-gluten anti-body recognized all protein fractions of molecular weight higher than 20 kDa. Quantitative ELISA analysis shows that the majority of samples has a gliadin-like protein content within the range of 80-260 mg/kg, excluding gluten-free flours and corresponding bakery products. Altogether, ß-glucan preparation has extremely high level of gliadin-like proteins. CONCLUSIONS: In the examined oat raw materials and foods the contents of immunoreactive amino acid sequences exceeded the limit of 20 mg/kg (considered as gluten-free) except for gluten-free flours (oat and  the prepared mixture) and the bakery products based on gluten-free flours. Unfortunately, the rest of oat raw materials and products cannot be considered gluten-free.

Trends and racial/ethnic disparities in gluten-sensitive problems in the United States: findings from the National Health and Nutrition Examination Surveys from 1988 to 2012.

OBJECTIVES: Racial disparities in the prevalence of celiac disease (CD) and the number of people without CD avoiding gluten (PWAG) in the United States are unknown. We aimed to describe racial differences in the prevalence of CD and PWAG, and evaluate the trends of CD in the noninstitutionalized civilian adult population of the US between 1988 and 2012. METHODS: A population-based cross-sectional study was conducted using data from the National Health and Nutrition Examination Surveys (NHANES) from 1988 to 1994, 1999 to 2004, and 2009 to 2012. Serum samples from the NHANES participants were tested for CD serology, which included IgA tissue transglutaminase (tTG IgA) and, if findings were abnormal, for IgA endomysial antibodies. Information about adherence to a gluten-free diet was obtained by means of an interviewer-administered questionnaire. RESULTS: In NHANES 2009-2012, the adjusted prevalence of CD was significantly higher (P<0.0001) among non-Hispanic whites (1.0%) than among non-Hispanic blacks (0.2%) and Hispanics (0.3%), whereas the adjusted prevalence of PWAG was significantly higher (P=0.01) in blacks (1.2%) as compared with Hispanics (0.5%) and whites (0.7%). The seroprevalence of CD in adults aged 50 years and older increased from 0.17% (95% confidence interval (CI) 0.03-0.33) in 1988-1994 to 0.44% (95% CI 0.24-0.81) in 2009-2012 (P<0.05). CONCLUSIONS: The overall prevalence of CD increased between 1988 and 2012 and is significantly more common in whites. In addition, a higher proportion of individuals maintaining a gluten-free diet in the absence of a diagnosis of CD are blacks.

Seroprevalence of celiac disease among United Arab Emirates healthy adult nationals: a gender disparity.

AIM: To determine celiac disease (CD) prevalence and associated manifestations or risk factors in healthy adult Emiratis. METHODS: It is a cross-sectional prospective study, recruiting 1197 (573 women and 624 men) healthy Emiratis between September 2007 and April 2008 among those who went to Al Ain Hospital to undertake the prenuptial examination. Test for anti-tissue transglutaminase (tTG) IgA antibodies was used for CD diagnosis. Subjects with positive results in the anti tTG antibodies assay were also tested for anti-endomysial (EMA) IgA antibodies. A structured interview was used to collect basic demographic and clinical recall data including: information on name, contact address, age, gender, education status, previous diagnosis of CD, diagnosis of CD in 1(st) degree relatives and history of "chronic diarrhea, anemia, headache, hepatitis, diabetes, tumor, and thyroid disorder". RESULTS: Fourteen blood samples (1.17%; 14/1197) were seropositive for CD. The latent CD seropositive patients were 13 women and 1 man and therefore the seroprevalence of CD was 1:86 (14/1197) for adult Emiratis: 1:44 (13/573) for women and 1:624 for men. Binary logistic regression revealed that history of chronic anemia (crude OR = 7.09; 95%CI: 2.32-21.61; P = 0.003) and being a woman (OR = 14.46; 95%CI: 1.89-110.91; P = 0.001) were associated with CD seropositivity. Whereas, the thyroid disorder showed a positive association with CD seropositivity that approach statistical significance (OR = 11.30; 95%CI: 1.32-96.95; P = 0.09) and therefore was included in the multiple logistic regression analysis, which showed that CD seropositivity is independently associated only with history of chronic anemia (OR = 4.58; 95%CI: 1.45-14.48; P = 0.01) and being a woman person (OR = 10.47; 95%CI: 1.33-82.14; P = 0.026). CONCLUSION: Compared to men the CD seroprevalence among women was remarkably higher. The CD association with women and chronic anemia is of importance from a public health perspective.

Serological assessment for celiac disease in IgA deficient adults.

PURPOSE: Selective immunoglobulin A deficiency is the most common primary immunodeficiency disorder that is strongly overrepresented among patients with celiac disease (CD). IgG antibodies against tissue transglutaminase (tTG) and deamidated gliadin peptides (DGP) serve as serological markers for CD in IgA deficient individuals, although the diagnostic value remains uncertain. The aim of this study was to investigate the prevalence of these markers in a large cohort of IgA deficient adults with confirmed or suspected CD and relate the findings to gluten free diet. METHODS: Sera from 488,156 individuals were screened for CD in seven Swedish clinical immunology laboratories between 1998 and 2012. In total, 356 out of 1,414 identified IgA deficient adults agreed to participate in this study and were resampled. Forty-seven IgA deficient blood donors served as controls. Analyses of IgG antibodies against tTG and DGP as well as HLA typing were performed and a questionnaire was used to investigate adherence to gluten free diet. Available biopsy results were collected. RESULTS: Out of the 356 IgA deficient resampled adults, 67 (18.8%) were positive for IgG anti-tTG and 79 (22.2%) for IgG anti-DGP, 54 had biopsy confirmed CD. Among the 47 IgA deficient blood donors, 4 (9%) were positive for IgG anti-tTG and 8 (17%) for anti-DGP. Four were diagnosed with biopsy verified CD, however, 2 of the patients were negative for all markers. Sixty-eight of 69 individuals with positive IgG anti-tTG were HLA-DQ2/DQ8 positive whereas 7 (18.9%) of the 37 individuals positive for IgG anti-DGP alone were not. CONCLUSIONS: IgG anti-tTG seems to be a more reliable marker for CD in IgA deficient adults whereas the diagnostic specificity of anti-DGP appears to be lower. High levels of IgG antibodies against tTG and DGP were frequently found in IgA deficient adults despite adhering to gluten free diet.

HLA DR-DQ genotyping by capillary electrophoresis for risk assessment for celiac disease.

The risk for celiac disease (CD) is clearly related to specific HLA DQA1 and DQB1 alleles, but HLA -typing is often considered too costly for frequent use.Here we present a method using sequence-specific primed PCR (PCR-SSP) for HLA-DR-DQ genotyping optimized for capillary electrophoresis on Applied Biosystems 3130xl Genetic Analyzer. Requiring a total of three PCR reactions and a single electrophoretic step, this method reduces the reagent expenses and technical time for directed HLA typing to distinguish risk alleles for CD, with a sufficient throughput for large-scale screening projects.

Gluten and gluten-free: issues and considerations of labeling regulations, detection methods, and assay validation.

Gluten is a commonly used cereal derivative found in bakery products, among other items. In some susceptible individuals, however, it triggers immune responses of different kinds; there is, to a lesser extent, the wheat allergy that is immunoglobulin E (IgE)-mediated and leads to histamine release and typical allergic symptoms. In this case, other water-soluble proteins, like albumins, are also involved. On the other hand, there is, more frequently, celiac disease (CD), where the gluten causes immune reactions in the intestines of certain individuals, leading to degeneration of villi, which typically leads to malabsorption of nutrients and, consequently, malnutrition. The only currently effective health strategy for affected consumers is avoidance of gluten-containing products, based on clear labeling rules. However, despite unanimously accepted Codex definitions by all member jurisdictions, the national implementation of equivalent laws shows significant differences. In the context of CD and in support of the gluten-free statement, regulatory enforcement, as well as manufacturers' quality controls are mostly based on analytical results. However, numerous methods are available, some of which have been validated better than others, and many provide different results on identical samples. Reasons include detection of different gluten components and variability in extraction efficiency due to different buffer compositions, especially from processed foods. Last but not least, the lack of reference materials is hindering the process of generating comparable data across different ELISA kits, as well as other methods. How can such data still be used to support a gluten-free claim? New methodologies, in particular mass spectrometric analysis of gluten derived peptides, are being introduced in numerous laboratories. This methodology is not only capable of detecting gluten derived peptides but can also differentiate between and quantitate wheat, barley, rye, and oat. This paper presents analytical limitations, as well as promising new approaches in support of industry and enforcement activities to ensure compliance with the gluten-free claim under the current regulatory framework.

Antibodies against deamidated gliadin peptides and tissue transglutaminase for diagnosis of pediatric celiac disease.

OBJECTIVES: The aim of the present study was to evaluate diagnostic performance and actual costs in clinical practice of immumoglobulin (Ig)G/IgA deamidated gliadin peptide antibodies (DGP) as a complement to IgA antibodies against tissue transglutaminase (tTG) for the diagnosis of pediatric celiac disease (CD). METHODS: All of the consecutive patients younger than 18 years tested for tTG and/or DGP, who underwent duodenal biopsy because of suspected CD in Stockholm and Gothenburg, Sweden, from 2008 to 2010, were included. Medical records were reviewed. RESULTS: Of 537 children who underwent duodenal biopsy, 278 (52%) had CD. A total of 71 (13%) were younger than 2 years and 16 (4%) had IgA deficiency. Sensitivity and specificity for tTG were 94% and 86%, respectively. Corresponding values for DGP were 91% and 26%. Positive predictive values (PPV) were 88% for tTG and 51% for DGP. There were 148 children who were tTG-negative and DGP-positive, of which only 5% (8/148) had villous atrophy. Among children younger than 2 years with normal IgA, PPV was 96% (25/26) for tTG and 48% (24/50) for DGP. In 16 IgA-deficient children, 11 were DGP positive, of which 5 had CD (PPV 45%). Eight of 278 cases of CD would possibly have been missed without DGP. The cost of adding DGP and consequently more biopsies to be able to detect 8 extra cases of CD was [Euro sign]399,520 or [Euro sign]49,940 per case. CONCLUSIONS: For diagnosing CD, tTG is superior to DGP, even in children younger than 2 years. Combining tTG and DGP does not provide a better tradeoff between number of missed cases of CD, number of unnecessary duodenal biopsies, and cost than tTG alone.