Comparison of printed glycan array, suspension array and ELISA in the detection of human anti-glycan antibodies

Anti-glycan antibodies represent a vast and yet insufficiently investigated subpopulation of naturally occurring and adaptive antibodies in humans. Recently, a variety of glycan-based microarrays emerged, allowing high-throughput profiling of a large repertoire of antibodies. As there are no direct approaches for comparison and evaluation of multi-glycan assays we compared three glycan-based immunoassays, namely printed glycan array (PGA), fluorescent microsphere-based suspension array (SA) and ELISA for their efficacy and selectivity in profiling anti-glycan antibodies in a cohort of 48 patients with and without ovarian cancer. The ABO blood group glycan antigens were selected as well recognized ligands for sensitivity and specificity assessments. As another ligand we selected P1, a member of the P blood group system recently identified by PGA as a potential ovarian cancer biomarker. All three glyco-immunoassays reflected the known ABO blood groups with high performance. In contrast, anti-P1 antibody binding profiles displayed much lower concordance. Whilst anti-P1 antibody levels between benign controls and ovarian cancer patients were significantly discriminated using PGA (p = 0.004), we got only similar results using SA (p = 0.03) but not for ELISA. Our findings demonstrate that whilst assays were largely positively correlated, each presents unique characteristic features and should be validated by an independent patient cohort rather than another array technique. The variety between methods presumably reflects the differences in glycan presentation and the antigen/antibody ratio, assay conditions and detection technique. This indicates that the glycan-antibody interaction of interest has to guide the assay selection

Systemic α-synuclein injection triggers selective neuronal pathology as seen in patients with Parkinson’s disease

Abstract: Parkinson’s disease (PD) is an α-synucleinopathy characterized by the progressive loss of specific neuronal populations. Here, we develop a novel approach to transvascularly deliver proteins of complex quaternary structures, including α-synuclein preformed fibrils (pff). We show that a single systemic administration of α-synuclein pff triggers pathological transformation of endogenous α-synuclein in non-transgenic rats, which leads to neurodegeneration in discrete brain regions. Specifically, pff-exposed animals displayed a progressive deterioration in gastrointestinal and olfactory functions, which corresponded with the presence of cellular pathology in the central and enteric nervous systems. The α-synuclein pathology generated was both time dependent and region specific. Interestingly, the most significant neuropathological changes were observed in those brain regions affected in the early stages of PD. Our data therefore demonstrate for the first time that a single, transvascular administration of α-synuclein pff can lead to selective regional neuropathology resembling the premotor stage of idiopathic PD. Furthermore, this novel delivery approach could also be used to deliver a range of other pathogenic, as well as therapeutic, protein cargos transvascularly to the brain

Mannose-based molecular patterns on stealth microspheres for receptor-specific targeting of human antigen-presenting cells

The targeting of antigen-presenting cells has recently gained strong attention for both targeted vaccine delivery and immunomodulation. We prepared surface-modified stealth microspheres that display various mannose-based ligands at graded ligand densities to target phagocytic C-type lectin receptors (CLRs) on human dendritic cells (DCs) and macrophages. Decoration of microspheres with carbohydrate ligands was achieved (i) by electrostatic surface assembly of mannan onto previously formed adlayers of poly(l-lysine) (PLL) or a mix of PLL and poly( l-lysine)- graft-poly(ethylene glycol) (PLL-PEG), or (ii) through assembly of PLL-PEG equipped with small substructure mannoside ligands, such as mono- and trimannose, as terminal substitution of the PEG chains. Microspheres carrying mannoside ligands were also studied in combination with an integrin-targeting RGD peptide ligand. Because of the presence of a mannan or PEG corona, such microspheres were protected against protein adsorption and opsonization, thus allowing the formation of specific ligand-receptor interactions. Mannoside density was the major factor for the phagocytosis of mannoside-decorated microspheres, although with limited efficiency. This strengthens the recent hypothesis by other authors that the mannose receptor (MR) only acts as a phagocytic receptor when in conjunction with yet unidentified partner receptor(s). Analysis of DC surface markers for maturation revealed that neither surface-assembled mannan nor mannoside-modified surfaces on the microspheres could stimulate DC maturation. Thus, phagocytosis upon recognition by CLRs alone cannot trigger DC activation toward a T helper response. The microparticulate platform established in this work represents a promising tool for systematic investigations of specific ligand-receptor interactions upon phagocytosis, including the screening for potential ligands and ligand combinations in the context of vaccine delivery and immunomodulation

Olfactory-induced brain activity in Parkinson's disease relates to the expression of event-related potentials : a functional magnetic resonance imaging study

Olfactory disorders are common in patients with idiopathic Parkinson's disease (IPD). In IPD patients with hyposmia olfactory event-related potentials (ERPs) are typically found to be delayed or absent. Altered ERPs in IPD patients may also be consistent with reduced neuronal activity in the medial temporal lobe following olfactory stimulation, as demonstrated by functional magnetic resonance imaging (fMRI). We analyzed ERPs and fMRI scans of hyposmic IPD patients (n=18) to gain further insight about the brain regions involved in generation of olfactory ERPs. Patients were separated into two groups (n=9 per group), based on the detectability (+) or non-detectability (-) of ERPs. Central activation during olfactory stimulation was examined using fMRI. Both ERP+ and ERP- patients showed activity in brain areas relevant to olfactory processing, such as the amygdala, parahippocampal regions, and temporal regions (BA 37, 21/22). Comparison of both groups revealed higher activation in ERP+ patients, especially in the amygdala, parahippocampal cortex, inferior frontal gyrus (BA 47), insula, cingulate gyrus, striatum, and inferior temporal gyrus. The relationship between the expression of olfactory ERPs and cortical activation patterns seen during olfactory stimulation in fMRI in IPD patients supports the idea that ERPs are a sensitive marker of neurodegeneration in olfactory regions. In accordance with current neuropathological staging concepts, olfactory ERPs may be reflecting pathological changes in olfactory regions, independent of the typically observed nigro-striatal degeneration in IPD. Reduced activation of primary olfactory areas in the ERP-group may reflect a severe disruption of olfactory processing in these patients