Fiche publication
Date publication
février 2016
Journal
Analytical and bioanalytical chemistry
Auteurs
Membres identifiés du Cancéropôle Est :
Dr CIANFERANI Sarah
Tous les auteurs :
Lanshoeft C, Wolf T, Heudi O, Cianférani S, Barteau S, Walles M, Picard F, Kretz O
Lien Pubmed
Résumé
In the present study, the application of a liquid chromatography high-resolution mass spectrometry (LC-HRMS) analytical assay for the quantitative analysis of a recombinant human immunoglobulin G1 (hIgG1) in rat serum is reported using three generic peptides GPSVFPLAPSSK (GPS), TTPPVLDSDGSFFLYSK (TTP), and VVSVLTVLHQDWLNGK (VVS). Moreover, the deamidation site of a fourth peptide FNWYVDGVEVHNAK (FNW) was identified and further excluded from the assay evaluation due to the inaccuracy of the quantitative results. The rat serum samples were spiked with a fully labeled hIgG1 as internal standard (ISTD). The digestion with trypsin was performed onto the pellet prior to peptide analysis by LC-HRMS using a quadrupole time of flight (QTOF) mass analyzer operating in selected reaction monitoring (SRM) mode with enhanced duty cycles (EDC). The assay linearity for the three investigated peptides was established for a hIgG1 (hIgG1A) from 1.00 to 1000 μg mL(-1) with a mean coefficient of determination (R (2)) higher than 0.9868. The inter-day accuracy and precision obtained in rat serum over 3 days were ≤11.4 and ≤10.5%, respectively. Short-term stability on the auto-sampler at 6 °C for 30 h, at RT for 48 h, and a 100-fold dilution factor were demonstrated. In addition, QC samples prepared in cynomolgus monkey serum and measured with the present method met the acceptance criteria of ±20.0 and ≤20.0% for all three peptides regarding accuracy and precision, respectively. The LC-HRMS method was applied to the analysis of samples from five individual cynomolgus monkeys dosed with a second hIgG1 (hIgG1B) and consistent data were obtained compared to the LC-MS/MS method (conventional triple quadrupole (QqQ) mass analyzer operating in SRM). The present data demonstrate that LC-HRMS can be used for the quantitative analysis of hIgG1 in both species and that quantification is not only limited to classical QqQ instruments.
Mots clés
Amino Acid Sequence, Animals, Chromatography, Liquid, methods, Female, Humans, Immunoglobulin G, blood, Macaca fascicularis, Mass Spectrometry, methods, Molecular Sequence Data, Peptides, chemistry, Rats, Recombinant Proteins, blood, Sensitivity and Specificity, Spectrometry, Mass, Electrospray Ionization, methods
Référence
Anal Bioanal Chem. 2016 Feb;408(6):1687-99
