In Vitro Performance of a Long-Range Surface Plasmon Hydrogel Aptasensor for Continuous and Real-Time Vancomycin Measurement in Human Serum.

This study investigated the suitability of surface modification for a long-range surface plasmon (LRSP) aptasensor using two different hydrogels, aiming at real-time monitoring of vancomycin (VCM) in undiluted serum and blood. Three different layer structures were formed on a gold surface of LRSP sensor chip using poly[2-methacryloyloxyethyl phosphorylcholine (MPC)-co-N-methacryloyl-(L)-tyrosinemethylester (MAT)] (PMM) and poly[MPC-co-2-ethylhexyl methacrylate (EHMA)-co-MAT] (PMEM). The peptide aptamer for VCM was immobilized in PMM and PMEM via MAT. Among four differently prepared sensor chips, the LRSP hydrogel aptasensor with PMM, referred to as the PMM hydrogel, exhibited the highest sensor output and superior antifouling properties. Following the optimization of the PMM hydrogel preparation conditions, the shelf life of the PMM hydrogel was determined to exceed 2 weeks, and the same sensor chip could be used for 102 days without significant performance deterioration. The PMM hydrogel was then applied for VCM measurement in undiluted serum in vitro, where it demonstrated a limit of detection of 0.098 µM and a dynamic range of 0.18-100 µM, covering the therapeutic range. Additionally, the PMM hydrogel enabled the continuous measurement of various VCM concentrations in serum without rinsing and showed a concentration-dependent output in undiluted blood. These findings underscore the potential of the PMM hydrogel for real-time and direct monitoring of VCM in body fluids.

In vitro SELEX and application of an African swine fever virus (ASFV) p30 protein specific aptamer.

The African swine fever virus (ASFV) has caused severe economic losses in the pig industry. To monitor ASFV spread, the p30 protein has been identified as an ideal infection marker due to its early and long-term expression during the ASFV infection period. Timely monitoring of ASFV p30 enables the detection of ASFV infection and assessment of disease progression. Aptamers are an outstanding substitute for antibodies to develop an efficient tool for ASFV p30 protein detection. In this study, a series of aptamer candidates were screened by in vitro magnetic bead-based systematic evolution of ligands by exponential enrichment (MB-SELEX). An aptamer (Atc-20) finally showed high specificity and affinity (Kd = 140 ± 10 pM) against ASFV p30 protein after truncation and affinity assessment. Furthermore, an aptamer/antibody heterogeneous sandwich detection assay was designed based on Atc20, achieving a linear detection of ASFV p30 ranging from 8 to 125 ng/ml and a detection limit (LOD) of 0.61 ng/ml. This assay showed good analytical performances and effectively detected p30 protein in diluted serum samples, presenting promising potential for the development of ASFV biosensors.

Biopanning the mimotopes of aflatoxin B1 and their immunogenicity / Panning de mimotopo aflatoxina B1 e sua imunogenicidade

Utilizando um anticorpo monoclonal contra a aflatoxina B1 (AFB1) como ligante, foi identificado um mimotopo específico de aflatoxina B1 após se realizarem quatro ciclos de seleção biológica de 7-peptídeos aleatórios em biblioteca de fago exibida. O mimotopo é denominado P10, e sua sequência de aminoácidos é YRRHEKD. O soro imunológico de ratos Balb/c imunizados com P10 foi especificamente ligado à aflatoxina B1-albumina, indicando que o anticorpo era específico ao AFB1. Esses resultados sugerem que é possível desenvolver a vacina baseada em mimotopo associado à toxina.(AU)

Los aptámeros como novedosa herramienta diagnóstica y terapéutica y su potencial uso en parasitología / Aptamers as a novel diagnostic and therapeutic tool and their potential use in parasitology

Los aptámeros son secuencias de ADN o ARN de cadena sencilla que adoptan la forma de estructuras tridimensionales únicas, lo cual les permite reconocer un blanco específico con gran afinidad. Sus usos potenciales abarcan, entre otros, el diagnóstico de enfermedades, el desarrollo de nuevos agentes terapéuticos, la detección de riesgos alimentarios, la producción de biosensores, la detección de toxinas, el transporte de fármacos en el organismo y la señalización de nanopartículas. El pegaptanib es el único aptámero aprobado para uso comercial por la Food and Drug Administration (FDA). Otros aptámeros para el tratamiento de enfermedades están en la fase clínica de desarrollo. En parasitología, se destacan los estudios que se vienen realizando en Leishmania spp., con la obtención de aptámeros que reconocen la proteína de unión a poliA (LiPABP) y que pueden tener potencial utilidad en la investigación, el diagnóstico y el tratamiento de la leishmaniasis. En cuanto a la malaria, se han obtenido aptámeros que permiten identificar eritrocitos infectados e inhiben la formación de rosetas, y otros que prometen ser alternativas para el diagnóstico al detectar de forma específica la proteína lactato deshidrogenasa (PfLDH). Para Cryptosporidium parvuum se han seleccionado aptámeros que detectan ooquistes a partir de alimentos o aguas contaminadas. Para Entamoeba histolytica se han aislado dos aptámeros llamados C4 y C5, que inhiben la proliferación in vitro de los trofozoítos y tienen potencial terapéutico. Los aptámeros contra Trypanosoma cruzi inhiben la invasión de células LLC-MK2 (de riñón de mono) en un 50 a 70 % y aquellos contra T. brucei transportan moléculas tóxicas al lisosoma parasitario como una novedosa estrategia terapéutica. Los datos recopilados en esta revisión destacan los aptámeros como una alternativa para la investigación, el diagnóstico y el tratamiento contra parásitos de interés nacional.

Aptamers are single-stranded DNA or RNA sequences that adopt unique three-dimensional structures that allow them to recognize a specific target with high affinity. They can potentially be used for the diagnosis of diseases, as new therapeutic agents, for the detection of food risks, as biosensors, for the detection of toxins, and as drug carriers and nanoparticle markers, among other applications. To date, an aptamer called pegaptanib is the only aptamer approved by the Food and Drug Administration (FDA) for commercial use. Other aptamers are in different clinical stages of development for the treatment of different diseases. In parasitology, investigations carried out with parasites such as Leishmania spp. allowed the acquisition of aptamers that recognize the polyA-binding protein LiPABP and may have potential applications in research and diagnosis and even as therapeutic agents. Regarding malaria, aptamers have been obtained that allow the identification of infected erythrocytes or inhibit the formation of rosettes, along with those that provide promising alternatives for diagnosis by specifically detecting the protein lactate dehydrogenase (PfLDH). In Cryptosporidium parvum allow the detection of oocysts in contaminated food or water. In Entamoeba histolytica, two aptamers called C4 and C5, which inhibit the proliferation of trophozoites in vitro and have potential use as therapeutic agents, have been isolated. Aptamers obtained against Trypanosoma cruzi inhibit the invasion of LLC-MK2 (from monkey kidney) cells by 50-70%, and in T. brucei, aptamers with the potential to transport toxic molecules to the parasitic lysosome were identified as a novel therapeutic strategy. The data collected in this review highlight aptamers as a novel alternative in the research, diagnosis, and treatment of parasites of national interest.