Master Degree / Yüksek Lisans Tezleri
Permanent URI for this collectionhttps://hdl.handle.net/11147/3008
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Master Thesis Effect of Gold Nanorod Properties on Lspr Response(01. Izmir Institute of Technology, 2023) Bulmuş Zareie, Volga; Tekin, Hüseyin Cumhur; Bulmuş Zareie, Esma Volga; Tekin, Hüseyin Cumhur; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyOptical qualities make gold nanorods (GNRs) excellent for plasmonic biosensors. Localized surface plasmon resonance (LSPR) phenomenon which occurs on GNR surfaces enables the creation of highly sensitive biosensors. The physical properties such as aspect ratio and size are directly related to the LSPR response of GNRs. The aim of this study is to investigate the impact of the aspect ratio (AR) and the interparticle distance on the localized surface plasmon resonance (LSPR) response of GNRs decorated glass sensor chips. For this aim, GNRs were first synthesized using a seed-mediated growth method. The effect of AgNO3 concentration on the AR of GNRs was investigated. It was observed that increasing AgNO3 concentration resulted in GNRs with higher AR and a red shift in the longitudinal plasmon peak wavelength. GNRs with an AR of 4, 6 and 8 were successfully synthesized. Next, the effect of the stabilizer molecule type and molecular weight on the distribution of GNRs on the silanized glass surface was investigated. It was found that the APTES modified glass surfaces cannot be coated with CTAB stabilized GNRs. Using GNRs stabilized with PEG5K resulted in a more homogeneous distribution of GNRs on the glass surface with respect to GNRs stabilized with PEG2K. The interparticle distance between GNRs on the glass surface was successfully controlled by simply concentrating or diluting the GNR solution used for coating the glass surfaces. It was observed that the LSPR peak shifts decreased upon binding of analytes as the interparticle distance between GNRs decreased in the studied range. On the other hand, as the AR decreased, the LSPR response of the GNRs shifted blue. The results presented in this thesis may contribute to future research to improve the potential of LSPR-based biosensors for diverse biomedical and diagnostic applications.Master Thesis Development of an Advanced Lspr-Based Biosensor Chip for Rapid Detection of Border Disease Virus(01. Izmir Institute of Technology, 2023) Bulmuş Zareie, Volga; Tekin, Hüseyin Cumhur; Bulmuş Zareie, Esma Volga; Tekin, Hüseyin Cumhur; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThe Border Disease Virus (BDV) is responsible for causing fetal deathly infection, leading to annual occurrences of affected farms. BDV, along with other pestiviruses such as classical swine fever virus (CSFV) and bovine viral diarrhea virus (BVDV), are known to cause major losses in stock farming. These losses can result in reproductive failure, expensive inspections, and other impacts on livestock health. The current detection methods of BDV include various techniques such as RT-PCR, ELISA, VNT, and immunofluorescence assays. These methods, although reliable, may require specialized equipment, time-consuming procedures, and laboratory facilities, making them less suitable for rapid on-site detection. Hence, it is imperative to employ diverse methodologies for detection of BDV. LSPR-based biosensors are a subset of plasmonic biosensors that exhibit numerous advantages for diverse applications. LSPR-based biosensors are particularly well-suited for the production of compact, practical devices for rapid, on-site detection of analytes. The aim of this study is to design and fabricate a biosensor chip utilizing LSPR technology for potential BDV detection. For this aim, glass surfaces were functionalized with gold nanorods modified with a BDV-specific primer sequence, complementary single-strand DNA sequence of 19 bases, and fabricated with PMMA microchannels. Different concentrations of target BDV-DNAsequence ranging from 0.01 pM to 100 nM were exposed to the channels, and the LSPR response was quantified using a Vis-NIR spectrometer. The limit of quantification of the biosensor chips was determined to be 10 pM, while the limit of detection was found to be less than or equal to 1 pM. The sensitivity of the biosensor chips was calculated to be 0.0567 nm/RIU. The dynamic range of the biochips lies between 10 pM to 100 pM.Master Thesis Enhancement of Bioavailability of Vitamin D by Nano-Sized Delivery Systems(01. Izmir Institute of Technology, 2023) Bulmuş Zareie, Volga; Kılıç Özdemir, Sevgi; Kılıç Özdemir, Sevgi; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03.02. Department of Chemical Engineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyStudies have indicated that Vitamin D (VitD) may decrease tumor invasiveness and propensity to metastasize. Cholecalciferol (VitD3) is the passive form of VitD3 and converts to active calcitriol through two-step hydroxylation reactions in the body, promoting binding to VitD-receptors (VDR). However, some breast cancer cells, especially MDA-MB-231, have very low levels of VDR. Besides, VitD3 suffers from first pass-effect of the liver which causes deactivation of VitD3. Therefore, new approaches are needed to increase VitD3 level in the cancerous sites. In this study, VitD3 was loaded into liposomes, which were subsequently coated by Fucoidan (FUC) to promote their binding to MDA-MB-231 cancer cells. Fucoidan strongly binds to P-selectins overexpressed in the breast cancer cells, blocking the cancer cells to adhere on the platelets to carry within the body, causing metastasis. Doxorubicin (DOX), being considered as the one of the most effective chemotherapeutic agents against breast cancer, was also loaded into liposomes in a similar manner. By liposomal encapsulations and fucoidan coating, it was aimed to deliver the all-cargo directly to the cancerous site and enhance the bioavailability of both agents at the target site. It was seen that liposomal VitD3 was more effective than free form to inhibit cell proliferation and, therapeutic potential of DOX increased with VitD3.VitD3 loaded FUC coated liposomes at optimized concentrations has a comparable effect with DOX-loaded liposomes with and without FUC coating. Overall, these results suggested that VitD3 and DOX loaded and FUC coated liposomes can be applied as combined therapy in cancer treatment.Master Thesis Design of Localized Surface Plasmon Resonance (lspr) Based Biosensor for Detecting a Potential Cancer Biomarker(Izmir Institute of Technology, 2020) Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyConventional methods for detection of cancer are invasive, expensive and not suitable for early diagnosis. Therefore, demand for simple, sensitive and rapid biosensors for detection of cancer have been enormous. Gold nanorods (GNRs) have been ideal materials for utilization in biosensors because of their exceptional optical properties. Localized surface plasmon resonance (LSPR) which is created on GNR surface can be used for the development of label-free and sensitive biosensor systems. LSPR responds to changes in the refractive index of the surroundings and this change can be observed as the shift in the maximum absorption wavelengths. In this thesis, an LSPR based GNR biosensor was developed for sensitive detection of a sialic acid as a potential cancer biomarker. For this purpose, GNRs were synthesized at around 40-50 nm in length. Afterwards, glass surfaces were coated with GNRs and functionalized with self-assembling molecules. Specific monoclonal antibodies(Ab) were conjugated to the surface. The surface modifications were characterized via contact angle, scanning electron microscope, Fourier transform infrared spectroscopy and zeta potential. Ab-functionalized glass surfaces were used to quantitatively detect specific molecular bindings via LSPR. The sensitivity of the biosensor was determined as 281 RIU/nm. The detection limit in PBS was 1 nM, while in serum it was found to be as 10 nM because of the high protein content of serum. Control experiments showed that the developed biosensor chip was selective. The proposed system is promising for early diagnosis of cancer since it can detect a potential cancer biomarker at concentrations as low as nanomolar level.Master Thesis Synthesis of Cationic Star Polymers Via Raft Polymerization(Izmir Institute of Technology, 2018) Dursun, Gürbüz; Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThe aim of this master thesis is to synthesize cleavable, core-crosslinked, cationic, new star polymers as potential siRNA carrier systems. The core-crosslinked star polymers of aminoethyl aminoethyl methacrylate (AEAEMA) monomer were synthesized for the first time in the literature. A crosslinker, potentially cleavable in cell cytosol, bis(2-methacryloyl) oxyethyl disulfide, and AEAEMA monomer with Boc-protection groups (BocAEAEMA) were polymerized using P(OEGMA) macroRAFT agents (5 kDa or 10 kDa). The polymerization kinetics revealed a linear increase in ([M]0/[M]) with increasing polymerization time, indicating the RAFT-controlled polymerization mechanism. The incorporation of arms into star structure completed between 2 and 8 hours of polymerization, leading to star formation yields of approx. 55%. Star polymers with narrow PDI values (between 1.52 and 1.80) and controllable molecular weights (between 116 kDa and 620 kDa) were synthesized using a P(OEGMA) macroRAFT agent of 10 kDa. Increasing crosslinker to macroRAFT agent ratio led to increase in crosslinker conversion and decrease in BocAEAEMA conversion. Increasing BocAEAEMA to macroRAFT agent ratio had the exact opposite effect. The use of a macroRAFT agent of 5 kDa led to a dramatic increase in molecular weight (up to 3370 kDa). The hydrodynamic sizes (Dh) of water soluble, cationic star polymers, obtained after deprotection of amino groups, dropped from 2-36 nm to 1-18 nm after treatment with a reducing agent in water, indicating the cleavable nature of the star structures. Among the star polymers tested, the polymer having 7 arms of POEGMA (10 kDa) and a degree of polymerization of AEAEMA (DPAEAEMA) of 70 efficiently complexed with siRNA at a nitrogen/phosphate ratio of 1. While this polymer showed higher cytotoxicity, when compared with other star polymers tested, all polymers were significantly less toxic, compared to branched PEI, a golden standard cationic polymer for siRNA delivery.Master Thesis Sericin-Polymer Conjugates: Preparation and Physicochemical Characterization(Izmir Institute of Technology, 2017) Gül, Abdulkadir; Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologySericin is a protein derived from silkworm, Bombyx mori, and has several useful properties as a natural biomaterial such as antioxidant character, moisturizing ability, hydrogel forming property and most importantly immunogenic inertness. The aim of this thesis is to prepare and physicochemically characterize sericin-polymer conjugates as potential natural-synthetic hybrid biomaterials with enhanced properties for drug delivery and tissue engineering applications. For this purpose, three polymers having the same degree of polymerization (n~42) and varying chemical nature, i.e. poly(oligoethylene glycol methacrylate), P(OEGMA) hydrophilic and neutral, poly(hydroxyethylmethacrylate) P(HEMA) less hydrophilic and neutral, and poly(dimethylaminoethyl methacrylate) P(DMAEMA) hydrophilic and cationic after quaternization, were first synthesized via reversible addition fragmentation chain transfer (RAFT) polymerization. Each polymer was characterized via nuclear magnetic resonance (1H-NMR) and gel permeation chromatography (GPC). Separately, molecular weight and isoelectric point of sericin were characterized using various techniques including Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and High-Resolution Two-Dimensional Polyacrylamide Gel Electrophoresis (2D-PAGE). Polymers were then covalently conjugated to sericin using NHS/EDC chemistry. The conjugates were characterized using SDS-PAGE, GPC and DLS (Dynamic Light Scattering). The SDS-PAGE and GPC results showed the successful preparation of the conjugates. DLS revealed that the hydrodynamic size of P(OEGMA) and P(DMAEMA) polymers and their conjugates were between 1 and 10 nm as they are soluble in PBS and do not form aggregates. Unlike the other two polymers, although the size of P(HEMA) polymer was observed to be 3.24 ± 0.62 nm, the DLS measurements of P(HEMA) conjugates indicated the presence of self-organization and aggregation of Sericin-P(HEMA) conjugates in aqeuous solution. Consequently, the size of sericin-P(HEMA) conjugates were found to be 530 ± 60.83 and 223.3 ± 25.2, respectively.Master Thesis Design and Construction of Portable Localized Surface Plasmon Resonance Device for Detection of Biological Molecules(Izmir Institute of Technology, 2017) Gül, Aytaç; Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyPoint-of-care devices giving rapid results in non-laboratory settings have become important for biosensor applicattions in a wide range of fields including medical, food, agriculture and pharmaceutical. This work aims to portable device based on localized surface plasmon resonance spectroscopy (LSPR-S) as a potential biosensor platform that can be used in non-laboratory settings for rapid detection of biological molecules at hifh sensivity. The thesis can be divided into two parts: In the first part, the design and consctruction of the device including both the mechanical and electronic parts are presented. The mechanical section includes the integration of the parts to build the device and microchannels designed with the aid of a three-dimensional drawing program Solid Works 2015. The second step of the construction process was the installation of electronic components onto the device. The electronicpart consists of a light source, fiber optic cables, a spectrometer and a temperature sensor. In conclusion, a portable LSPR-S device with an integrated microchannel system has been produced, which potentially allows analysing low volumes of sample without the need to label the molecules. The second part of the thesis covers the studies towards the preparation and application of sensing platforms for the LSPR-S device constructed to enable the rapid detection of biplogical molecules at high sensitivity. These included the preparation of gold nanorods and nanoparticles-based LSPR-S detection of model antibody-antigen and bacteria-bacteriophage interactions, respectively. Studies conducted in this section have led to the conclusion that LSPR-S-based biosensor platforms developed in thisis are promising solutions to overcome current challenges in biosensor applications.Master Thesis Interactions of Cancer Cells and Macrophages on the Egf-Egfr Axis: Chemotaxis, Haptotaxis or Direct Contact?(Izmir Institute of Technology, 2017) Önal, Sevgi; Pesen Okvur, Devrim; Bulmuş Zareie, Volga; Pesen Okvur, Devrim; Bulmuş Zareie, Esma Volga; 04.03. Department of Molecular Biology and Genetics; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 04. Faculty of Science; 01. Izmir Institute of TechnologyBreast cancer cells (BCC) and macrophages are known to interact via epidermal growth factor (EGF) produced by macrophages and colony stimulating factor-1 (CSF-1) produced by BCC. Despite contradictory findings, this interaction is perceived as a paracrine loop. Yet, the underlying mechanism of interaction remains unclear. Here, we investigated interactions of BCC with macrophages in 2D and 3D. BCC did not show chemotaxis to macrophages in custom designed 3D cell-on-a-chip devices, which was in agreement with ELISA results showing that macrophage-derived-EGF was not secreted into macrophage-conditioned-medium. Live cell imaging of BCC in the presence and absence of iressa showed that macrophages but not macrophage-derivedmatrix modulated adhesion and motility of BCC in 2D. 3D co-culture experiments in matrigel and collagen showed that BCC changed their multicellular organization in the presence of macrophages. In custom designed 3D co-culture cell-on-a-chip devices, macrophages reduced and promoted migration of BCC in matrigel and collagen, respectively. Furthermore, adherent but not suspended BCC endocytosed EGFR when in contact with macrophages. Collectively, our data revealed that macrophages showed chemotaxis towards BCC-derived-CSF-1 whereas BCC required direct contact to interact with macrophage-derived-EGF. We propose that the interaction between cancer cells and macrophages is a paracrine-juxtacrine loop of CSF-1 and EGF, respectively.Master Thesis In Vitro Evaluation of Poly (2-((2 Amino) Ethyl Methacrylate) as a Potential Sirna Delivery Agent(Izmir Institute of Technology, 2015) Seyrantepe, Volkan; Seyrantepe, Volkan; Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; Seyrantepe, Volkan; 03.01. Department of Bioengineering; 04.03. Department of Molecular Biology and Genetics; 03. Faculty of Engineering; 04. Faculty of Science; 01. Izmir Institute of TechnologyThe aim of this thesis is to investigate poly(2-((2-aminoethyl)amino)ethyl methacrylate) (P(AEAEMA)) as a potential siRNA carrier. For this purpose, an amine containing monomer 2-((tert-butoxycarbonyl) (2-((tert-butoxy carbonyl) amino) ethyl) amino) ethyl methacrylate (BocAEAEMA) was synthesized. Reversible addition-fragmentation chain transfer (RAFT) polymerization was performed to prepare homo- and block co-polymers of BocAEAEMA. The synthesized polymers -P(AEAEMA)19, P(AEAEMA)41 and P(PEGMA)12-b-P(AEAEMA)32- were characterized via NMR and GPC. The cytotoxicity of the polymers was investigated in vitro using ovarian cancer cell line (Skov-3-luc) via MTT assay. The polymers did not show any toxic effect on cells in 24 h. The ability of the BocAEAEMA polymers to form polyplexes with siRNA was investigated via gel electrophoresis. P(AEAEMA)19, P(AEAEMA)41 and P(PEGMA)12-b-P(AEAEMA)32 could efficiently form complexes with siRNA at an N/P ratio of 5, 2, and 2 respectively. Gel electrophoresis analysis revealed that P(AEAEMA)41 and P(PEGMA)12-b-P(AEAEMA)32 could protect siRNA against serum components at least for 6 h. Block copolymer, when complexed with siRNA at an N/P ratio of 10, could protect siRNA longer (24 h) when compared with the homopolymer. The size and surface charge of the polyplexes were investigated by DLS. The diameter of the P(AEAEMA)41-siRNA complexes was found to be lower than 110 nm at all N/P ratios tested. In contrast, P(PEGMA)12-b-P(AEAEMA)32-siRNA complexes (except the complex prepared at the N/P ratio of 2), displayed aggregation tendency. All polyplexes displayed positive zeta potential. The zeta potential of the homopolymer was found to be higher than the copolymer at the N/P ratio of 2. Finally, in order to determine siRNA transfection ability of the polymers, luciferase assay was optimized using a commercial transfection reagent lipofectamine RNAimax. The optimized assay conditions will be used in future studies to determine the transfection efficiency of the polymers.Master Thesis Development of Arginine-Containing Well-Defined Polymers(Izmir Institute of Technology, 2014) Taykoz, Damla; Bulmuş Zareie, Volga; Bulmuş Zareie, Esma Volga; 03.01. Department of Bioengineering; 03. Faculty of Engineering; 01. Izmir Institute of TechnologyThe aim of this work is to synthesize arginine-containing well-defined polymers via reversible addition-fragmentation chain transfer (RAFT) polymerization and perform preliminary investigation on the use of these polymers in nucleic acid complexation for potential gene therapy applications. Pentafluorophenyl methacrylate (PFMA) was chosen as an active ester monomer to produce polymers having functional groups available for further modification. RAFT polymerization of PFMA was performed varying polymerization conditions such as feed composition and polymerization temperature. Polymers (PPFMA) were characterized using nuclear magnetic resonance (NMR) spectroscopy and gel permeation chromatography. Linear increase in ln[M]0/[M] with polymerization time, and number average molecular weight (Mn) with monomer conversion indicated RAFT controlled polymerization of PFMA under the conditions tested. Furthermore, block copolymers of PFMA with poly(ethylene glycol) methacrylate (PEGMA) as a biocompatible component were prepared. Copolymerization was studied using both P(PFMA) and P(PEGMA) as macro RAFT agent. Copolymerization kinetic studies indicated that chain extension block copolymerizations were successfully performed using both macroRAFT agents. P(PFMA) was reacted with arginine methylester (AME) in the presence of triethylamine (TEA). 100% of P(PFMA) active ester groups could be modified with AME at a polymer/AME/TEA mole ratio of 1/1/3, as determined by 1H-NMR spectroscopy. The AME modified polymers were complexed with a 681-bp DNA fragment through electrostatic interactions at varying nitrogen/phosphate (N/P) ratios. Gel electrophoresis experiments revealed that AME-modified P(PFMA) was able to complex with DNA at a N/P ratio of 200. Furthermore, the hydrodynamic diameter (Dh) of polymer/DNA complexes in phosphate buffer saline was found to be 58 nm, while the free DNA displayed a Dh of 109 nm, indicating the complexation of DNA by AME-modified P(PFMA).