title: A novel method for serum lipoprotein profiling using high performance capillary isotachophoresis.
abstract: A new capillary isotachophoresis (cITP) method for lipoprotein profiling with superior lipoprotein coverage compared to previous methods has been developed, resolving twice as many lipoprotein species (18 peaks/fractions) in serum or plasma in less than 9.5 min. For this, a novel mixture of 24 spacers, including amino acids, dipeptides and sulfonic acids, was developed and fine-tuned, using predictive software (PeakMaster) and testing of spiked serum samples. Lipoprotein peaks were identified by serum-spiking with reference lipoproteins. Compatibility with common lipophilic stains for selective lipoprotein detection with either UV/Vis or laser-induced fluorescence was demonstrated. A special new capillary with a neutral coating (combining water-compatible OV1701-OH deactivation and methylation) was used for the first time for electrodriven separations, allowing very stable separations in a pH 8.8-9.4 gradient system, being functional for more than 100 injections. Excellent reproducibility was achieved, with coefficients of variation lower than 2.6% for absolute migration times. Comparison was performed with human plasma samples analyzed by NMR, leading to similar results with cITP after multivariate statistics, regarding group-clustering and lipoprotein species correlation. The new cITP method was applied to the analysis of serum samples from a LDL receptor knock-out mice model fed either a normal diet or a western-type diet. Differences in the lipoprotein levels and in the sublipoprotein types were detected, showing a shift to more atherogenic particles due to the high cholesterol diet. In summary, this novel method will allow more detailed and informative profiling of lipoprotein particle subtypes for cardiovascular disease research.
authors: Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Faculty of Science, Universiteit Leiden, Einsteinweg 55, 2300 RA Leiden, The Netherlands. Electronic address: emorenog@ucm.es.
authors: Estefanía
authors: E
authors: Moreno-Gordaliza
authors: Department of Epidemiology, Erasmus Medical Centre, PO Box 2040, 3000 CA Rotterdam, The Netherlands.
authors: Sven J
authors: SJ
authors: van der Lee
authors: Department of Epidemiology, Erasmus Medical Centre, PO Box 2040, 3000 CA Rotterdam, The Netherlands.
authors: Ayşe
authors: A
authors: Demirkan
authors: Department of Epidemiology, Erasmus Medical Centre, PO Box 2040, 3000 CA Rotterdam, The Netherlands.
authors: Cornelia M
authors: CM
authors: van Duijn
authors: Division of Biopharmaceutics, Leiden Academic Centre for Drug Research, Faculty of Science, Universiteit Leiden, Einsteinweg 55, 2300 RA Leiden, The Netherlands.
authors: Johan
authors: J
authors: Kuiper
authors: Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Faculty of Science, Universiteit Leiden, Einsteinweg 55, 2300 RA Leiden, The Netherlands.
authors: Petrus W
authors: PW
authors: Lindenburg
authors: Division of Analytical Biosciences, Leiden Academic Centre for Drug Research, Faculty of Science, Universiteit Leiden, Einsteinweg 55, 2300 RA Leiden, The Netherlands.
authors: Thomas
authors: T
authors: Hankemeier
mesh_terms: D000818:Animals; D001774:Blood Chemical Analysis / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D055786:Gene Knockout Techniques; D006801:Humans; D057625:Isotachophoresis / Q000379:methods*; D008074:Lipoproteins / Q000097:blood* / Q000302:isolation & purification*; D051379:Mice; D011973:Receptors, LDL / Q000172:deficiency / Q000235:genetics; D012984:Software
chemical_list: D008074:Lipoproteins; D011973:Receptors, LDL
keywords: Atherosclerosis; Capillary coating; Capillary isotachophoresis; Hypercholesterolemia; Lipoproteins; Spacers