NNF Center for Basic Metabolic Research, Department of Biomedical Sciences, Faculty of Health Sciences.
Eur J Endocrinol. 2014 Mar 8;170(4):529-38. doi: 10.1530/EJE-13-0941. Print 2014 Apr.
To determine the specificity and sensitivity of assays carried out using commercially available kits for glucagon and/or oxyntomodulin measurements.
Ten different assay kits used for the measurement of either glucagon or oxyntomodulin concentrations were obtained. Solutions of synthetic glucagon (proglucagon (PG) residues 3361), oxyntomodulin (PG residues 3369) and glicentin (PG residues 169) were prepared and peptide concentrations were verified by quantitative amino acid analysis and a processing-independent in-house RIA. Peptides were added to the matrix (assay buffer) supplied with the kits (concentration range: 1.25-300 pmol/l) and to human plasma and recoveries were determined. Assays yielding meaningful results were analysed for precision and sensitivity by repeated analysis and ability to discriminate low concentrations.
Three assays were specific for glucagon (carried out using the Millipore (Billerica, MA, USA), Bio-Rad (Sundbyberg, Sweden), and ALPCO (Salem, NH, USA) and Yanaihara Institute (Shizuoka, Japan) kits), but none was specific for oxyntomodulin. The assay carried out using the Phoenix (Burlingame, CA, USA) glucagon kit measured the concentrations of all three peptides (total glucagon) equally. Sensitivity and precision were generally poor; the assay carried out using the Millipore RIA kit performed best with a sensitivity around 10 pmol/l. Assays carried out using the BlueGene (Shanghai, China), USCN LIFE (Wuhan, China) (oxyntomodulin and glucagon), MyBioSource (San Diego, CA, USA) and Phoenix oxyntomodulin kits yielded inconsistent results.
确定使用市售试剂盒进行胰高血糖素和/或胰高血糖素样肽-1 测量的特异性和灵敏度。
获得了用于测量胰高血糖素或胰高血糖素样肽-1 浓度的 10 种不同的测定试剂盒。制备了合成胰高血糖素（前胰高血糖素（PG）残基 3361）、胰高血糖素样肽-1（PG 残基 3369）和糖依赖性胰岛素释放肽（PG 残基 169）的溶液，并通过定量氨基酸分析和独立于加工的内部 RIA 验证了肽浓度。将肽添加到试剂盒提供的基质（测定缓冲液）中（浓度范围：1.25-300pmol/l）并添加到人血浆中，然后确定回收率。通过重复分析和区分低浓度的能力，对产生有意义结果的测定进行精密度和灵敏度分析。
有 3 种测定方法对胰高血糖素具有特异性（使用 Millipore（比勒陀利亚，MA，美国）、Bio-Rad（桑德比贝里，瑞典）和 ALPCO（塞勒姆，NH，美国）和矢野研究所（静冈，日本）试剂盒进行），但没有一种方法对胰高血糖素样肽-1 具有特异性。使用 Phoenix（伯林盖姆，CA，美国）胰高血糖素试剂盒进行的测定同样测量了所有三种肽（总胰高血糖素）的浓度。灵敏度和精密度通常较差；使用 Millipore RIA 试剂盒进行的测定具有约 10pmol/l 的灵敏度，效果最好。使用 BlueGene（上海，中国）、USCN LIFE（武汉，中国）（胰高血糖素和胰高血糖素样肽-1）、MyBioSource（圣地亚哥，CA，美国）和 Phoenix 胰高血糖素样肽-1 试剂盒进行的测定得出的结果不一致。
