Rabia Cakir-Koc

Production of anti-SAG1 IgY antibody against Toxoplasma gondii parasites and evaluation of antibody activity by ELISA method

Chicken egg yolk antibody, also known as immunoglobulin Y (IgY), is the predominant class of serum immunoglobulin in birds. IgY has many advantages over mammalian antibodies, such as enhanced immunogenicity conserved mammalian proteins exhibited in birds due to their phylogenetic distance, non-invasive rapid, and economical collection system. However, there are limited studies about IgY production against Toxoplasma, which is a worldwide veterinary and public health problem. In this study, the production of specific IgY antibodies against the surface antigen 1 (SAG1) protein of Toxoplasma gondii and the determination of antibody activity via the enzyme-linked immunosorbent assay (ELISA) method were conducted. According to ELISA, Western blot, and NanoDrop results, specific and higher amounts of IgY antibody against SAG1 were obtained with this study. Considering the advantages of IgY and importance of SAG1 for the diagnosis of toxoplasmosis, it is expected that anti-SAG1 IgY will play an increasing role and gain commercial value in research, diagnostics, and immunotherapy against toxoplasmosis in the future.

Utility of the Microculture Method in Non-Invasive Samples Obtained from an Experimental Murine Model with Asymptomatic Leishmaniasis

The sensitivity of diagnostic methods for visceral leishmaniasis (VL) decreases because of the low number of parasites and antibody amounts in asymptomatic healthy donors who are not suitable for invasive sample acquisition procedures. Therefore, new studies are urgently needed to improve the sensitivity and specificity of the diagnostic approaches in non-invasive samples. In this study, the sensitivity of the microculture method (MCM) was compared with polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and immunofluorescent antibody test (IFAT) methods in an experimental murine model with asymptomatic leishmaniasis. Results showed that the percent of positive samples in ELISA, IFAT, and peripheral blood (PB) -PCR tests were 17.64%, 8.82%, and 5.88%, respectively, whereas 100% positive results were obtained with MCM and MCM-PCR methods. Thus, this study, for the first time, showed that MCM is more sensitive, specific, and economic than other methods, and the sensitivity of PCR that was performed to samples obtained from MCM was higher than sensitivity of the PCR method sampled by PB.

Novel NAC-loaded poly(lactide-co-glycolide acid) nanoparticles for cataract treatment: preparation, characterization, evaluation of structure, cytotoxicity, and molecular docking studies

Background N-acetylcarnosine (NAC), a dipeptide with powerful antioxidant properties that is extensively used as a pharmaceutical prodrug for the treatment of cataract and acute gastric disease, was investigated by molecular dynamics with the GROMACS program in order to understand the solvent effect on peptide conformation of the peptide molecule used as a component of a drug and which presents substantial information on where drug molecules bind and how they exert their effects. Besides, molecular docking simulation was performed by using the AutoDock Vina program which identify the kind of interaction between the drug and proteins. A delivery system based on poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with NAC (NAC-PLGA-NPs) for the treatment of cataract was prepared for the first time in this study in order to enhance drug bioavailability and biocompatibility. The objective of this work was to prepare and evaluate the structural formulation, characterization, and cytotoxicity studies of NAC-loaded NPs based on PLGA for cataract treatment. Methods PLGA and NAC-loaded PLGA NPs were prepared using the double emulsion (w/o/w) method, and characterizations of the NPs were carried out with UV–Vis spectrometer to determine drug concentration, the Zeta-sizer system to analyze size and zeta potential, FTIR spectrometer to determine the incorporation of drug and PLGA, and TEM analysis for morphological evaluation. Results NAC-loaded PLGA NPs were successfully obtained according to UV–Vis and FTIR spectroscopy, Zeta-sizer system. And it was clearly observed from the TEM analysis that the peptide-loaded NPs had spherical and non-aggregated morphology. Also, the NPs had low toxicity at lower concentrations, and toxicity was augmented by increasing the concentration of the drug. Discussion The NAC molecule, which has been investigated as a drug molecule due to its antioxidant and oxidative stress-reducing properties, especially in cataract treatment, was encapsulated with a PLGA polymer in order to increase drug bioavailability. This study may contribute to the design of drugs for cataract treatment with better reactivity and stability.

A Global Problem of Toxoplasmosis

Abstract The causative agent of toxoplasmosis, Toxoplasma gondii , is an intracellular parasite and member of the Apicomplexan family. Toxoplasmosis can be seen all over the world together with a growing human population. People can be infected with T. gondii in several ways, such as contact with infected animals, contaminated blood, a leukocytes transfusion, congenital transmission, and foodborne transmission. There is a special risk for pregnant women who get infected with T. gondii , as the fetus may be stillborn or have some abnormalities such as hydrocephalus, intracranial calcifications, microcephaly, fetal growth retardation, ascites, or hepatosplenomegaly. Ocular toxoplasmosis causes blindness and loss of eyes. Toxoplasmosis becomes especially important regarding nervous system infections. In this chapter, we describe the general characteristics and life cycle of T. gondii , epidemiology and transmission, diagnosis and treatment of toxoplasmosis, and clinical pictures of diseases. We especially focus on the relation between toxoplasmosis and the central nervous system.

Green synthesis of silver nanoparticles using Thymbra spicata L. var. spicata (zahter) aqueous leaf extract and evaluation of their morphology-dependent antibacterial and cytotoxic activity

Abstract Silver (Ag) nanoparticles (NPs) were green synthesized at room temperature using different concentrations of the Thymbra spicata L. var. spicata (Zahter) aqueous leaf extracts for the first time. With the synthesis of AgNPs using the leaf extract of Cynara scolymus (Artichoke) and Mentha piperita (Peppermint), the biological activities of the nanoparticles synthesized using leaf extract of three economically significant plants have been studied comparatively. Nanoparticles were characterized by different spectroscopic and microscopic analysis. TEM analysis of the biosynthesized AgNPs revealed that the size and shape of the AgNPs were changed with the plant extract concentration. Biologically synthesized AgNPs from leaf extracts of the three different plants displayed significant differences in antibacterial activity against two different gram-negative and gram-positive bacteria. Also, the results from this study show the shape dependence of the antibacterial and cytotoxic activity of silver nanoparticles synthesized using T. spicata leaf extract. The nanoparticles with different shapes exhibited the strongest antibacterial and cytotoxic activity compared to mostly spherical nanoparticles. Present results clearly indicate that biological activities of silver nanoparticles were affected by nanoparticle shape and the source of the plant extract used in the synthesis.

Influenza Diagnosis with a Specific Emphasis on the M2e Antigen as a Diagnostic Tool

The therapy, observation, inclusiveness, and preclusion of related diseases all influence the diagnosis of influenza. Particularly, the pandemic duration and diagnosis time for influenza are extremely important. After the appearance of symptoms, antiviral medication must be initiated within 48 h. Cell culture, real‐time polymerase chain reaction (PCR), flow cytometry, direct and indirect immunofluorescence methods, and the quick diagnosis test are all valuable approaches for the diagnosis of influenza. Different instruments, different time durations for the results, and different specialists characterize all these approaches. Antigen selection is of critical importance with regard to the specificity and sensitivity of these methods, especially the serological and rapid diagnosis tests. M2e, the highly conserved external domain of the influenza A M2 protein, is a potential differential diagnostic marker for influenza virus infection. This chapter reviews the studies that use M2e as a diagnosis agent, and it illuminates the role and importance of M2e in the diagnosis of influenza.

Molecular docking of immunogenic peptide of Toxoplasma gondii and encapsulation with polymer as vaccine candidate

Abstract Toxoplasma gondii is one of the most widely spread parasitic organisms in the world. T. gondii causes primary, chronic infection and mortality. Major surface antigen 1 is the most abundant tachyzoite surface protein and highly conserved between species and causes strong humoural response. Some studies showed that the peptide sequence of surface antigen has immunity. Therefore, tachyzoite surface antigenic peptide sequence is one of the good candidates for vaccine development. However, conformational information and delivery systems are very important parameters for vaccine development. Computational chemistry which is used as an effective method to perform drug or vaccine design provides important information on structure–activity relationship, biological effects of functional groups, molecular geometry, design of enzyme inhibitors and antagonists. The interaction of immunological peptides with protein systems was carried out by means of computing the free energy of binding using the molecular docking technique. Due to the major histocompatibility complex (MHC), proteins play a substantial role for adaptive immunity, the crystal structure of a MHC class I, which plays a pivotal role in the adaptive branch of the immune system, was preferred for docking calculations. A delivery system based on poly(lactic-co-glycolic acid) (PLGA) nanoparticles and peptide loaded PLGA nanoparticles was prepared in this study to improve the bioavailability of tachyzoite surface antigenic peptide sequence. Double emulsion method (water-in-oil-in-water or w/o/w) was used for synthesis of PLGA and peptide loaded PLGA nanoparticles. The average particle size, polydispersity index and zeta potential values of PLGA and peptide loaded PLGA nanoparticles were measured with zeta-sizer by using dynamic light scattering (DLS) technique. The scanning electron microscope (SEM) (Zeiss Supra 50 V) was used for imagining the peptide loaded PLGA nanoparticles. Cell toxicity of nanoparticles was assayed on AGS (gastric adenocarcinoma) cell line. To evaluate mitochondrial activity of cells and toxicity studies, XTT methods were carried out. In this study, we aimed to obtain specific immunological peptide loaded PLGA nanoparticles and characterize the formation with FTIR, zeta sizer and SEM imaging, and evaluate cytotoxicity and carry out molecular docking calculations of peptide–MHC protein in order to enlight in vivo events as vaccine candidate against T. gondii.

11 – Evaluation techniques

Characterizing structure and surface detail of nanoparticles at molecular or atomic level are essential for understanding their properties and their functional and technical performances in technological applications. The number of studies in literature body on manufacture and characterization of nanomaterials increase every day, since their area of utilization expands and their importance is appreciated more. Several techniques have been developed to directly or indirectly characterize nanomaterials. Some of these techniques are surface-sensitive, while others are element-specific. Evaluation techniques for structural analysis and biomechanical properties of the nanomaterials include advanced microscopy technologies, cellular studies with biomaterials, stem cell-based nanomaterial studies, and in vitro and in vivo assessments. For evaluation of nanomaterials, a single technique alone cannot provide enough information. Complementary techniques are required to understand the changes in nanomaterials during physical, chemical, or biochemical processing. Thus, this section covers the most common and advanced techniques for assessment of nanomaterial properties.