Search results for "Ultracentrifuge"
showing 10 items of 20 documents
Isolation and Analysis of Fasciola hepatica Extracellular Vesicles
2020
The finding of extracellular vesicles (EVs) as important players in parasite-parasite and host-parasite communications has led to an increasing number of reports in the literature. Different protocols have been developed for isolation and further characterization of EVs from parasitic helminths. In this chapter, we describe step by step procedures to isolate EVs secreted by Fasciola hepatica adults in culture, which could be also applied for other developmental stages of the parasite, as well as EVs present in plasma and urine. Along with classical isolation methods like differential ultracentrifugation, and more recent techniques like size exclusion chromatography (SEC), here we also refer…
Optimized production and purification of Coxsackievirus B1 vaccine and its preclinical evaluation in a mouse model.
2017
Coxsackie B viruses are among the most common enteroviruses, causing a wide range of diseases. Recent studies have also suggested that they may contribute to the development of type 1 diabetes. Vaccination would provide an effective way to prevent CVB infections, and the objective of this study was to develop an efficient vaccine production protocol for the generation of novel CVB vaccines. Various steps in the production of a formalin-inactivated Coxsackievirus B1 (CVB1) vaccine were optimized including the Multiplicity Of Infection (MOI) used for virus amplification, virus cultivation time, type of cell growth medium, virus purification method and formulation of the purified virus. Safety…
Molecular basis of a new type of C1q-deficiency associated with a non-functional low molecular weight (LMW) C1q: parallels and differences to other k…
1998
Analysis of an abnormal C1q molecule of individuals of a Moroccan family by ultracentrifugation in sucrose gradients revealed a low molecular weight C1q (LMW-C1q). We investigated the molecular basis of this defect by sequencing all six exons of the three C1q genes. One point mutation in the codon for Gly at position 15 (GGT) of the B chain was found resulting in an amino acid substitution to Asp (GAT). The exchange not only leads to an interruption of the collagen-like motif Gly-X-Y, but also introduces one negatively charged residue per B chain which results in two additional charges per structural subunit (A-B, C-C, A-B). The mutation which has been identified by DNA-sequencing in the C1…
Enhanced oxidative susceptibility and reduced antioxidant content of metabolic precursors of small, dense low-density lipoproteins.
2001
Abstract PURPOSE: Elevated plasma concentrations of low-density lipoproteins (LDL) increase risk for coronary heart disease. However, lipoprotein profiles rich in small, dense LDL particles confer greater risk than those that mainly consist of large, buoyant LDL. This may be due, in part, to the greater oxidative susceptibility of small, dense LDL. In the current studies, we tested whether differences in the oxidative behavior of buoyant and dense LDL arise from differences in their immediate metabolic precursors, intermediate-density lipoproteins. SUBJECTS AND METHODS: We compared the properties of intermediate-density lipoproteins and buoyant and dense LDL subfractions in 9 subjects with …
Purification and analysis of polyhistidine-tagged human parvovirus B19 VP1 and VP2 expressed in insect cells
2008
Human parvovirus B19 is an autonomously replicating human pathogen with a specific tropism for human erythroid progenitor cells. There is an interest in producing empty nucleocapsids of B19 as they can be used as tools in molecular biology and diagnostics. Native B19 virus particles are formed from two structural viral proteins, VP1 and VP2. The VP2 protein alone is able to self assemble and consequently form virus-like particles (VLPs) in heterologous expression systems. Purification of recombinant VLPs has been conducted using various traditional methods. These include laborious and time-consuming, e.g. cesium chloride or sucrose gradient ultracentrifugation steps, allowing limited workin…
Comparison of quantum dot-binding protein tags: Affinity determination by ultracentrifugation and FRET
2013
Abstract Background Hybrid complexes of proteins and colloidal semiconductor nanocrystals (quantum dots, QDs) are of increasing interest in various fields of biochemistry and biomedicine, for instance for biolabeling or drug transport. The usefulness of protein–QD complexes for such applications is dependent on the binding specificity and strength of the components. Often the binding properties of these components are difficult and time consuming to assess. Methods In this work we characterized the interaction between recombinant light harvesting chlorophyll a / b complex (LHCII) and CdTe/CdSe/ZnS QDs by using ultracentrifugation and fluorescence resonance energy transfer (FRET) assay exper…
Preparation of Native and Recombinant Light-Harvesting Chlorophyll-<I>a/b </I>Complex
2004
Procedures to isolate native light-harvesting chlorophyll-a/b complex (LHCIIb) and to reconstitute recombinant LHCIIb are described. Separation of trimeric from monomeric forms and free pigment by sucrose density-gradient ultracentrifugation can be applied to both native and reconstituted complexes. The preparations are characterized by their pigment composition, protein pattern, and spectral properties.
A Classical Synthesis of the Collagen-like Peptides with the Sequence Z(GlyProPro)nOBut and their characterization with circular dichroism and ultrac…
1975
Oligopeptides with the sequence Z(GlyProPro)nOBut (n = 3–7) were synthesized along a clasical pathway. Generally long fragments were condensed in order to allow an easy separation of the products from the reactants. It is established by molecular weight determination and measurements of circular dichroism that the peptides form the collagen-like triple helix in methanol and that they assume the random coil conformation in dilute acetic acid. The circular dichroism spectra agree reasonably well with the corresponding spectra for (ProProGly)10 which was obtained by solid phase synthesis in another laboratory.
Numerical analysis of density gradient centrifugation profiles from eukaryotic DNA
1990
A numerical method for the deconvolution of superimposed Gaussian distributions with a unique solution has been proposed by Medgyessy [10]. We have tested the usefulness of this method for the analysis of density gradient centrifugation profiles from eukaryotic DNA, which are normally composed from overlapping Gaussian distributed profiles of several subcomponents with different mean buoyant densities. From the analysis of human DNA and from model calculations we conclude that major subcomponents can be identified by this method, if they differ in their buoyant density by approximatly 0.005 g/ml. Minor components can only be identified if the total DNA has been fractionated according to buo…