Biarc Jordane, Simon Romain, Fonbonne Catherine, Léonard Jean-François, Gautier Jean-Charles, Pasquier Olivier, Lemoine Jerôme, Salvador Arnaud
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France.
Sanofi, 13, Quai Jules Guesde, 94400 Vitry-sur-Seine, France.
J Chromatogr A. 2015 Jun 5;1397:81-5. doi: 10.1016/j.chroma.2015.04.003. Epub 2015 Apr 8.
Podocalyxin is a protein present in specialized glomerulus cells called podocytes and may be released in the urine in case of kidney injury. In this context, its quantification could be of great interest in order to monitor glomerular injury. Liquid chromatography tandem mass spectrometry (LC-MS/MS), in selected reaction monitoring (SRM) mode, has been demonstrated as a powerful technique that can be applied to protein quantification. This paper describes the development of a quantification method of human podocalyxin in urine by LC-MS/MS in SRM mode by monitoring one proteotypic peptide with an isotope-dilution standardization strategy employing (13)C/(15)N labelled peptides. Inter/intra assay precisions and accuracies of the assay were below 10% and between 90% and 106.1%, respectively. In addition, the method was linear between 0.78 and 100 ng/mL and could therefore be used to quantify endogenous level of podocalyxin that was estimated between 15.2 and 44.2 ng/mL in urine samples from healthy donor.
足细胞毒素是一种存在于称为足细胞的特殊肾小球细胞中的蛋白质，在肾脏损伤时可能会释放到尿液中。在这种情况下，对其进行定量分析对于监测肾小球损伤可能具有重要意义。液相色谱串联质谱法（LC-MS/MS）在选择反应监测（SRM）模式下，已被证明是一种可用于蛋白质定量分析的强大技术。本文描述了一种通过LC-MS/MS在SRM模式下，采用（13）C/（15）N标记肽的同位素稀释标准化策略，监测一条蛋白质特征肽来定量尿液中人类足细胞毒素的方法。该分析方法的批间/批内精密度和准确度分别低于10%以及在90%至106.1%之间。此外，该方法在0.78至100 ng/mL之间呈线性，因此可用于定量健康供体尿液样本中估计在15.2至44.2 ng/mL之间的足细胞毒素内源性水平。
