Gluten in Beers: Evaluation of Reproducibility of the R5-Based Competitive Enzyme-Linked Immunosorbent Assay Method Using Real Samples

Maria del Pilar Fernandez-Gil, Marian Bustamante, Jon Esparta, Olaia Martínez, J. Miranda, E. Simón
{"title":"Gluten in Beers: Evaluation of Reproducibility of the R5-Based Competitive Enzyme-Linked Immunosorbent Assay Method Using Real Samples","authors":"Maria del Pilar Fernandez-Gil, Marian Bustamante, Jon Esparta, Olaia Martínez, J. Miranda, E. Simón","doi":"10.3390/proceedings2023091354","DOIUrl":null,"url":null,"abstract":". Abstract: Beer is the most widely consumed alcoholic beverage in Europe. In many occasions, its consumption is linked to social relations and a fruitive use. To comply with this, the market should offer gluten-free beers that are safe to be consumed by people with celiac disease or those who need to avoid gluten. Brewing hydrolyzes gluten, and this compels the analytical determination of this hydrolyzed protein to be carried out using a competitive ELISA method. The most commonly used competitive ELISA for this purpose is based on the R5 antibody, which has some disadvantages, such as less robustness compared to the homologous sandwich ELISA. The aim of this study was to evaluate the reproducibility of the R5-based competitive ELISA through detecting gluten in beers that intended to achieve a gluten-free label. Thirty-seven samples of beers in which gluten was detected (range 10–80 mg/kg gluten) were analyzed under intermediate precision conditions (e.g., different days and different analysts). Each sample was analyzed 3–20 times. A total of 185 tests were performed and statistically analyzed. The mean calculation of the relative standard deviation (RSD) has a median of 13.6% (range 2.1–23.4%). The samples were pooled according to their gluten content (expressed as mg/kg or ppm gluten) and the median for each interval as follows: beers containing 10–20 ppm (n = 9): RSD 16.1% (range 2.7–19.9%); 21-40 ppm (n = 20): RSD 12.7% (range 2.8–21.5%); and 41–140 ppm (n = 8): RSD 13.7% (range 2.1–23.4%). The main variability in precision was found in the samples with a low gluten content, close to the limit of quantification. This could be due to the fact that small differences in the measured absorbances in this range make a significant difference in quantification. Our results suggest that the precision of the assayed method in our laboratory was satisfactory, in line with the expectable results of other ELISA methods. An internal reproducibility of 20% could be a reliable limit for any testing laboratory. Without evaluating other factors such as accuracy, the data findings point to an elevated uncertainty value for this analytical method.","PeriodicalId":507355,"journal":{"name":"The 14th European Nutrition Conference FENS 2023","volume":"13 12","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2024-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"The 14th European Nutrition Conference FENS 2023","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.3390/proceedings2023091354","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

Abstract

. Abstract: Beer is the most widely consumed alcoholic beverage in Europe. In many occasions, its consumption is linked to social relations and a fruitive use. To comply with this, the market should offer gluten-free beers that are safe to be consumed by people with celiac disease or those who need to avoid gluten. Brewing hydrolyzes gluten, and this compels the analytical determination of this hydrolyzed protein to be carried out using a competitive ELISA method. The most commonly used competitive ELISA for this purpose is based on the R5 antibody, which has some disadvantages, such as less robustness compared to the homologous sandwich ELISA. The aim of this study was to evaluate the reproducibility of the R5-based competitive ELISA through detecting gluten in beers that intended to achieve a gluten-free label. Thirty-seven samples of beers in which gluten was detected (range 10–80 mg/kg gluten) were analyzed under intermediate precision conditions (e.g., different days and different analysts). Each sample was analyzed 3–20 times. A total of 185 tests were performed and statistically analyzed. The mean calculation of the relative standard deviation (RSD) has a median of 13.6% (range 2.1–23.4%). The samples were pooled according to their gluten content (expressed as mg/kg or ppm gluten) and the median for each interval as follows: beers containing 10–20 ppm (n = 9): RSD 16.1% (range 2.7–19.9%); 21-40 ppm (n = 20): RSD 12.7% (range 2.8–21.5%); and 41–140 ppm (n = 8): RSD 13.7% (range 2.1–23.4%). The main variability in precision was found in the samples with a low gluten content, close to the limit of quantification. This could be due to the fact that small differences in the measured absorbances in this range make a significant difference in quantification. Our results suggest that the precision of the assayed method in our laboratory was satisfactory, in line with the expectable results of other ELISA methods. An internal reproducibility of 20% could be a reliable limit for any testing laboratory. Without evaluating other factors such as accuracy, the data findings point to an elevated uncertainty value for this analytical method.
啤酒中的麸质使用真实样品评估基于 R5 的竞争性酶联免疫吸附测定法的重现性
.摘要:啤酒是欧洲消费最广泛的酒精饮料。在许多场合,啤酒消费与社交关系和果味用途有关。为此,市场上应该提供不含麸质的啤酒,让乳糜泻患者或需要避免食用麸质的人可以安全饮用。酿造过程会水解麸质蛋白,因此必须使用竞争性 ELISA 方法对水解蛋白进行分析测定。最常用的竞争性酶联免疫吸附法是基于 R5 抗体的,它有一些缺点,如与同源的夹心酶联免疫吸附法相比稳定性较差。本研究的目的是通过检测无麸质标签啤酒中的麸质,评估基于 R5 的竞争性 ELISA 的重现性。在中等精度条件下(如不同日期和不同分析师)分析了 37 个检测到麸质(范围 10-80 mg/kg gluten)的啤酒样品。每个样品分析 3-20 次。总共进行了 185 次测试并进行了统计分析。相对标准偏差 (RSD) 的平均计算中值为 13.6%(范围为 2.1-23.4%)。根据麸质含量(以毫克/千克或百万分之麸质表示)和每个区间的中位数对样品进行了汇总,具体如下:麸质含量为 10-20 ppm 的啤酒(n = 9):RSD 16.1%(范围 2.7-19.9%);21-40 ppm(n = 20):RSD 12.7%(范围 2.8-21.5%);41-140 ppm(n = 8):RSD 13.7%(范围 2.1-23.4%)。精度的主要变化出现在麸质含量较低的样品中,接近定量限。这可能是由于在此范围内测量的吸光度的微小差异会对定量产生重大影响。我们的结果表明,我们实验室的检测方法精度令人满意,与其他 ELISA 方法的预期结果一致。对于任何检测实验室来说,20% 的内部重现性都是一个可靠的限度。在不评估准确度等其他因素的情况下,数据结果表明这种分析方法的不确定值较高。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
自引率
0.00%
发文量
0
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信