ABSTRACT
Several diagnostic tools have been developed for clinical and epidemiological assays. RT-PCR and antigen detection tests are more useful for diagnosis of acute disease, while antibody tests allow the estimation of exposure in the population. Currently, there is an urgent need for the development of diagnostic tests for COVID-19 that can be used for large-scale epidemiological sampling. Through a comprehensive strategy, potential 16 mer antigenic peptides suited for antibody-based SARS-CoV-2 diagnosis were identified. A systematic scan of the three structural proteins (S,N and M) and the non-structural proteins (ORFs) present in the SARS-CoV-2 virus was conducted through the combination of immunoinformatic methods, peptide SPOT synthesis and an immunoassay with cellulose-bound peptides (Pepscan). The Pepscan filter paper sheets with synthetic peptides were tested against pools of sera of COVID-19 patients. Antibody recognition showed a strong signal for peptides corresponding to the S, N and M proteins of SARS-CoV-2 virus, but not for the ORFs proteins. The peptides exhibiting higher signal intensity were found in the C-terminal region of the N protein. Several peptides of this region showed strong recognition with all three immunoglobulins in the pools of sera. The differential reactivity observed between the different immunoglobulin isotypes (IgA, IgM and IgG) within different regions of the S and N proteins, can be advantageous for ensuring accurate diagnosis of all infected patients, with different times of exposure to infection. Few peptides of the M protein showed antibody recognition and no recognition was observed for peptides of the ORFs proteins.
Subject(s)
COVID-19 Serological Testing/methods , Coronavirus M Proteins/immunology , Coronavirus Nucleocapsid Proteins/immunology , Informatics/methods , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Viral/blood , Computational Biology , Coronavirus M Proteins/genetics , Coronavirus Nucleocapsid Proteins/genetics , Epitope Mapping , Epitopes, B-Lymphocyte/genetics , Humans , Immunoglobulin A/blood , Immunoglobulin G/blood , Immunoglobulin M/blood , Peptides/genetics , Spike Glycoprotein, Coronavirus/geneticsABSTRACT
The cAMP-dependent protein kinase (PKA) is the best understood member of the superfamily of serine-threonine protein kinases and is involved in controlling a variety of cellular processes. Measurements of PKA activity traditionally relied on the use of [(32)P]-labeled ATP as the phosphate donor and a protein or peptide substrate as the phosphoaceptor. Recently non-isotopic assays for the PKA have been developed and this paper presents an improvement of a fluorometric assay for measuring the activity of PKA. Three peptides were synthesized with the following sequences: LRRASLG (Kemptide), LRRASLGK (Kemptide-Lys8) and LRRASLGGGLRRASLG (Bis-Kemptide), these have in common the substrate sequence recognized by the PKA (RRXS/TΨ), where X is any amino acid and Ψ is a hydrophobic amino acid. Optimal conditions were established for the non-radioactive assay to detect the PKA activity by phosphorylation of these three peptides that are covalently linked to fluorescamine at their N-terminus. The phosphorylated and non-phosphorylated peptides were easily separated by electrophoresis, identified and quantified with optical densitometry and ultraviolet light. The fluorescamine-labeled Kemptide-Lys8 substrate (Fluram-Kemptide-Lys8) was used to calculate the Km and Vmax of the catalytic subunit of PKA from pig heart and showed a detection limit of 260 pmol, a linear range between 700 and 1150 pmol with a linear regression R (2) = 0.956.
Subject(s)
Cyclic AMP-Dependent Protein Kinases/chemistry , Enzyme Assays/methods , Fluorescamine/chemistry , Oligopeptides/chemistry , Amino Acid Sequence , Animals , Catalytic Domain , Cyclic AMP-Dependent Protein Kinases/isolation & purification , Kinetics , Myocardium/chemistry , Oligopeptides/chemical synthesis , Phosphorylation , Substrate Specificity , SwineABSTRACT
A group of substituted 1-benzyl-3-carbethoxy-2-quinolones, whose design was based on the structures of well established antimitotics, were synthesized by a three reactions sequence. Although the yields of the first reaction (condensation of o-nitrobenzaldehydes with diethyl malonate) could not be improved, the synthetic route turned out quite efficient.
Subject(s)
Antineoplastic Agents/chemical synthesis , Quinolones/chemical synthesis , Antineoplastic Agents/chemistry , Quinolones/chemistryABSTRACT
Un grupo de 1-bencil-3-carbethoxi-2-quinolonas sustituidas, diseñadas en base a las estructuras de antimitóticos bien establecidos, fueron sintetizados mediante una secuencia de tres reacciones. Aunque los rendimientos de la primera reacción (condensación de o-nitrobenzaldehídos sustituidos con malonato de dietilo) no pudieron ser mejorados, la ruta sintética resultó bastante eficiente
Subject(s)
Quinolones , Antineoplastic Agents , Quinolones , Antineoplastic AgentsABSTRACT
Amoxicilina es un antibiótico betalactámico bactericida perteneciente al grupo de las penicilinas semisintéticas con un amplio espectro de acción , es estructuralmente análogo a otros antibióticos de este grupo, pero la Amoxicilina se caracteriza por su absorción rápida y casi completa lo que garantiza unos niveles sanguíneos elevados y persistentes con la posibilidad de reducir convenientemente la frecuencia de la administración. Se describe el desarrollo de una tecnología para la producción industrial en forma de polvo para suspensión oral en la dosis de 125 mg para su uso pediátrico. Se utilizó la vía seca como alternativa para la elaboración del granulado , midiendo los parámetros físicos y químicos a lo largo de todo el estudio acorde con la USP XXII . Se realizó el estudio de estabilidad por dos años, logrando un producto con la calidad requerida para el uso a que está destinado. (AU)
Subject(s)
Amoxicillin , Lactams , Particulate MatterABSTRACT
Las cefalosporinas, se clasifican dentro de los antibióticos B-lactamicos de amplio espectro y su uso esta indicado en el tratamiento de un extenso conjunto de infecciones por gérmenes gram positivos y gram negativos. Estos actúan inhibiendo la síntesis de la pared celular de las bacterias, poseen una vida media larga lo que resulta beneficioso a la hora de su administración. Nuestro trabajo describirá el desarrollo de las formulaciones para la obtención de productos inyectables con la calidad requerida, así como el estudio de estabilidad de las cefalosporinas a fin de determinar la vida útil de las mismas, tanto en forma de polvo como después de reconstituidas. Los productos logrados cumplen los requerimientos más actuales de la farmacopea USP XXII. Con la introducción de estos antibióticos al mercado nacional, se logró sustituir en gran medida las cantidades importadas de estos productos y aumentar el surtido en los hospitales, de forma tal, que la población disponga de antibióticos eficaces e insustituibles fundamentalmente en las salas de terapia intensiva (AU)
