Polyethylene imine-based receptor immobilization for label free bioassays

Polyethylene imine (PEI) based immobilization of antibodies is described and the concept is proved on the label free assay of C-Reactive Protein (CRP). This novel immobilization method is composed of a hyperbranched PEI layer which was deposited at a high pH (9.5) on the sensor surface. The free amino groups of PEI were derivatized with neutravidin by Biotin N-hydroxysuccinimide ester and the biotinylated anti-CRP antibody immobilized on this layer. Direct binding assay of recombinant CRP was successfully performed in the low μg/ml concentrations using a label free optical waveguide biosensor

Cerebrospinal fluid neopterin as marker of the meningo-encephalitic stage of Trypanosoma brucei gambiense sleeping sickness.

BACKGROUND: Sleeping sickness, or human African trypanosomiasis (HAT), is a protozoan disease that affects rural communities in sub-Saharan Africa. Determination of the disease stage, essential for correct treatment, represents a key issue in the management of patients. In the present study we evaluated the potential of CXCL10, CXCL13, ICAM-1, VCAM-1, MMP-9, B2MG, neopterin and IgM to complement current methods for staging Trypanosoma brucei gambiense patients. METHODS AND FINDINGS: Five hundred and twelve T. b. gambiense HAT patients originated from Angola, Chad and the Democratic Republic of the Congo (D.R.C.). Their classification as stage 2 (S2) was based on the number of white blood cells (WBC) (>5/µL) or presence of parasites in the cerebrospinal fluid (CSF). The CSF concentration of the eight markers was first measured on a training cohort encompassing 100 patients (44 S1 and 56 S2). IgM and neopterin were the best in discriminating between the two stages of disease with 86.4% and 84.1% specificity respectively, at 100% sensitivity. When a validation cohort (412 patients) was tested, neopterin (14.3 nmol/L) correctly classified 88% of S1 and S2 patients, confirming its high staging power. On this second cohort, neopterin also predicted both the presence of parasites, and of neurological signs, with the same ability as IgM and WBC, the current reference for staging. CONCLUSIONS: This study has demonstrated that neopterin is an excellent biomarker for staging T. b. gambiense HAT patients. A rapid diagnostic test for detecting this metabolite in CSF could help in more accurate stage determination

Actigraphy in Human African Trypanosomiasis as a Tool for Objective Clinical Evaluation and Monitoring: A Pilot Study

The clinical picture of the parasitic disease human African trypanosomiasis (HAT, also called sleeping sickness) is dominated by sleep alterations. We here used actigraphy to evaluate patients affected by the Gambiense form of HAT. Actigraphy is based on the use of battery-run, wrist-worn devices similar to watches, widely used in middle-high income countries for ambulatory monitoring of sleep disturbances. This pilot study was motivated by the fact that the use of polysomnography, which is the gold standard technology for the evaluation of sleep disorders and has greatly contributed to the objective identification of signs of disease in HAT, faces tangible challenges in resource-limited countries where the disease is endemic. We here show that actigraphy provides objective data on the severity of sleep-wake disturbances that characterize HAT. This technique, which does not disturb the patient's routine activities and can be applied at home, could therefore represent an interesting, non-invasive tool for objective HAT clinical assessment and long-term monitoring under field conditions. The use of this method could provide an adjunct marker of HAT severity and for treatment follow-up, or be evaluated in combination with other disease biomarkers in body fluids that are currently under investigation in many laboratories

Ca2+ signals critical for egress and gametogenesis in malaria parasites depend on a multipass membrane protein that interacts with PKG.

Calcium signaling regulated by the cGMP-dependent protein kinase (PKG) controls key life cycle transitions in the malaria parasite. However, how calcium is mobilized from intracellular stores in the absence of canonical calcium channels in Plasmodium is unknown. Here, we identify a multipass membrane protein, ICM1, with homology to transporters and calcium channels that is tightly associated with PKG in both asexual blood stages and transmission stages. Phosphoproteomic analyses reveal multiple ICM1 phosphorylation events dependent on PKG activity. Stage-specific depletion of Plasmodium berghei ICM1 prevents gametogenesis due to a block in intracellular calcium mobilization, while conditional loss of Plasmodium falciparum ICM1 is detrimental for the parasite resulting in severely reduced calcium mobilization, defective egress, and lack of invasion. Our findings suggest that ICM1 is a key missing link in transducing PKG-dependent signals and provide previously unknown insights into atypical calcium homeostasis in malaria parasites essential for pathology and disease transmission

A study of the cambial zone and conductive phloem of common beech (Fagus sylvatica L.) using an image analysis method. I. Influence of tree age on the structure

Quantification is a major problem when using histology to study the influence of ecological factors on tree structure. This paper presents a method to prepare and to analyse transverse sections of cambial zone and of conductive phloem in bark samples. The following paper (II) presents the automated measurement procedure. Part I here describes and discusses the preparation method, and the influence of tree age on the observed structure. Highly contrasted images of samples extracted at breast height during dormancy were analysed with an automatic image analyser. Between three young (38 years) and three old (147 years) trees, age-related differences were identified by size and shape parameters, at both cell and tissue levels. In the cambial zone, older trees had larger and more rectangular fusiform initials. In the phloem, sieve tubes were also larger, but their shape did not change and the area for sap conduction was similar in both categories. Nevertheless, alterations were limited, and demanded statistical analysis to be identified and ascertained. The physiological implications of the structural changes are discussed

General characteristics of the 2 cohorts.

<p>NA: not available.</p>▵<p>: All patients were collected within the first 3 h after symptom onset.</p