Absolute quantification of podocalyxin, a potential biomarker of glomerular injury in human urine, by liquid chromatography-mass spectrometry.
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.
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France.
Jordane
J
Biarc
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France.
Romain
R
Simon
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France.
Catherine
C
Fonbonne
Sanofi, 13, Quai Jules Guesde, 94400 Vitry-sur-Seine, France.
Jean-François
JF
Léonard
Sanofi, 13, Quai Jules Guesde, 94400 Vitry-sur-Seine, France.
Jean-Charles
JC
Gautier
Sanofi, 13, Quai Jules Guesde, 94400 Vitry-sur-Seine, France.
Olivier
O
Pasquier
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France.
Jerôme
J
Lemoine
UMR 5280, Institut des Sciences Analytiques, Université de Lyon, Lyon 1, 5 Rue de la Doua, 69100 Villeurbanne, France. Electronic address: arnaud.salvador@univ-lyon1.fr.
Arnaud
A
Salvador
D058186:Acute Kidney Injury / Q000652:urine*; D015415:Biomarkers / Q000652:urine*; D002853:Chromatography, Liquid; D006801:Humans; D007201:Indicator Dilution Techniques; D057230:Limit of Detection; D015203:Reproducibility of Results; D012795:Sialoglycoproteins / Q000032:analysis*; D053719:Tandem Mass Spectrometry; D016482:Urinalysis / Q000379:methods*
D015415:Biomarkers; D012795:Sialoglycoproteins; C001643:podocalyxin
Biomarker; Podocalyxin; Protein; Quantification; SRM; Urine