Standardized peptidome profiling of human cerebrospinal fluid by magnetic bead separation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Peptidome profiling of human cerebrospinal fluid (CSF) is a promising tool to identify novel disease-associated biomarkers. Our aim was to develop a standardized protocol for reproducible peptidome profiling of CSF using magnetic bead (MB) separation followed by MALDI-TOF MS. Peptidome fractionation and profiling of CSF were performed using MBs with different surface functionalities. We investigated exogenous variables (storage conditions, freeze-thaw-cycles) and endogenous interferences (albumin, immunoglobulin, blood, leukocytes) in pooled CSF samples. We detected approximately 500 signals with an S/N ratio >10 and an overlap frequency of about 40% in non-pathological CSF. Within- and between-day imprecisions in relative signal intensities ranged from 3 to 28% and 7 to 47%, respectively. CSF storage at room temperature for up to 6 h and at 4 degrees C for up to 3 days did not significantly influence the mass spectra. Consecutive freeze-thaw-cycles significantly affected the mass spectra. High albumin and immunoglobulin content altered the CSF preparation using MB-HIC C8 beads. Blood contamination showed no effect on mass spectra up to a hemoglobin concentration of 0.075 micromol/L. The presence of leukocytes up to a cell number of 30 Mpt/L did not affect mass spectra. Our reliable pretreatment protocol allows standardization of preanalytical modalities and thereby enables reproducible peptidome profiling of human CSF using MB separation followed by MALDI-TOF MS.

Labeling of human neural precursor cells using ferromagnetic nanoparticles.

Fetal human neural precursor cells (NPCs) are unique with respect to their capacity to proliferate and to preserve their potential to differentiate into neurons and glia. Human mesencephalic neural precursor cells (hmNPCs) provide a source for dopaminergic neurons. Preclinical and clinical research will benefit from reliable in vivo tracking of transplanted cells. Here, we investigate the potency of very small superparamagnetic iron oxide particles (VSOPs) to label hmNPCs, the effect of VSOPs on survival, proliferation, and differentiation of hmNPCs, and the sensitivity of 1.5T magnetic resonance imaging (MRI) to detect labeled cells in living rats following transplantation. When incubated with VSOPs at 1.5 mM, >95% of hmNPCs incorporated VSOPs without detectable impact on cell viability (>90%) or proliferative capacity, as measured by the expression of proliferating cell nuclear antigen (PCNA) and cell cycle distribution. Labeled hmNPCs differentiate into neurons (>30%) and glia with no detectable difference compared to nonlabeled cells. Following transplantation into rat striata, marked paramagnetic signal changes were detected for as long as three months postsurgery using MRI, corresponding to the histologically-identified graft. Our data indicate that hmNPCs can be labeled with VSOPs without impairment of viability, proliferation, or multipotency. Labeled, transplanted cells are detectable in vivo using 1.5T MRI.