Aptamers are oligonucleotides that have the characteristic of recognizing a target with high affinity and specificity. Based on our previous studies, the aptamer probe Sgc8-c-Alexa647 is a promising tool for molecular imaging of PTK7, which is an interesting biomarker in cancer. In order to improve the delivery of this probe as well as create a novel drug delivery nanosystem targeted to the PTK7 receptor, we evaluate the co-association between the probe and preformed nanostructures. In this work, preformed pegylated liposomes (PPL) and linear and branched pristine polymeric micelles (PMs), based on PEO–PPO–PEO triblock copolymers were used: poloxamer F127^® and poloxamines T1307^® and T908^®. For it, Sgc8-c-Alexa647 and its co-association with the different nanostructures was exhaustively analyzed. DLS analysis showed nanometric sizes, and TEM and AFM showed notable differences between free- and co-associated probe. Likewise, all nanosystems were evaluated on A20 lymphoma cell line overexpressing PTK7, and the confocal microscopy images showed distinctness in cellular uptake. Finally, the biodistribution in BALB/c mice bearing lymphoma-tumor and pharmacokinetic study revealed an encouraging profile for T908-probe. All data obtained from this work suggested that PMs and, more specifically T908 ones, are good candidates to improve the pharmacokinetics and the tumor uptake of aptamer-based probes.

The aim of the present work is to prepare and characterize a functionalized latex with chloromethyl groups on the surface and to perform the covalent coupling of anti-human serum albumin (a-HSA) IgG protein. The chloromethyl-styrene latex (CMS) was synthesized by means of a core-shell emulsion polymerization in a batch reactor. The monodisperse-obtained latex was characterized by determining the diameter (TEM and PCS), the surface charge density (conductometric and potentiometric titration), the amount of chloromethyl groups on the surface (hydrolysis reaction), and the stability vs electrolyte concentration (turbidity measurements). Electrokinetic characterization was also performed (electrophoretic mobility versus pH and ionic strength). IgG was chemically bound to the latex particles under different sensitization and block-stabilization conditions. Colloidal stability of complexes was studied to select an immunolatex suitable for the development of latex immunoassays. The final part of this work consists of a study of the immunoreactivity of the IgG-latex complexes at different pH and ionic strength, in particular under physiological conditions. The results show that chemically bound IgG to chloromethyl latex provides an IgG-latex complex suitable for application in immunodiagnosis tests.

The aim of the present work is to study the influence of the sequential or competitive adsorption of anti-HSA (Human Serum Albumin) IgG and RSA (Rabbit Serum Albumin) onto a negatively charged polystyrene latex on the colloidal stability of the immunoiatex (IgG-RSA/latex). Anionic (1-2) latex was prepared by surfactant free emulsion polymerization. Adsorption of IgG and RSA on the latex 1-2 was performed at low ionic strength and different pHs. Sequential and competitive adsorption of a-HSA IgG and RSA were performed Colloidal stability of immunolatex was improved by RSA adsorption, especially in the competitive adsorption. In this last case, the immunological agglutination reactions of physically adsorbed IgG molecules onto anionic particles have been investigated by means of optical absorbance measurements. These measurements have been conducted under different experimental conditions to determine the most influential factors. Surface IgG and BSA densities, particle concentration in the reaction medium, and polyethylene glycol concentration are some of these factors. An amount of HSA lesser than 60 ng/ml has been possible to detect using this particle enhanced optical immunoassay.

Ischemic cerebrovascular accident (iCVA) is a public health issue, whose subjacent events involve the development of nitroxidative distress. Identifying biomarkers that assist in the diagnosis of this disease has clinically relevant implications. The aim of this study was to develop an analytic tool for measuring nitroxidative distress biomarkers, intended for application in clinical practice to enhance patient healthcare. Three enzyme linked immunosorbent assays (ELISA) were developed, with different detection objectives. One of them, in a sandwich format, quantifies the amount of fibrinogen in human plasma, an important glycoprotein involved in the blood coagulation process, contributing to thrombus formation and thereby participating in the mechanism of ischemic stroke. Another ELISA, also in a sandwich format, detects the presence of nitrotyrosine residues in fibrinogen from human plasma, a nitroxidative posttranslational modification resulting from the attack of peroxynitrite by-products on tyrosine residues present in proteins. The third one, in inhibition format, determines human plasma nitrotyrosine total content and was used to analyze human plasma samples from control and iCVA patients. Those two groups of plasma samples were analyzed using inhibition ELISA, revealing statistically significant differences in their nitrotyrosine content and molar ratios of nitrotyrosine to fibrinogen, which were higher in the iCVA group. This study provides evidence that nitroxidative distress occurs in ischemic stroke, as indicated by the detection of the biomarker nitrotyrosine. This finding supports other studies that also identified nitrotyrosine in ischemic stroke, through several different methods. This specific ELISA method is applicable for the rapid analysis of clinical samples, making it a potential clinical tool for assessing iCVA patients.