Development of au nanoparticles by the method of ultrasonic spray pyrolysis for the detection of antibodies against SARS-CoV-2

V doktorskem delu so predstavljene študije, eksperimentalno delo, tehnike karakterizacije, rezultati in analize, s pomočjo katerih smo potrdili hipotezo, da je možno Au nanodelce (AuND) pripravljene z metodo ultrazvočne razpršilne pirolize (USP), konjugirati s ciljnimi antigeni SARS-CoV-2 in rekombinantnim IgG, da bodo delovali v imunoserološkem hitrem testu na lateralni tok (LFIA), za potrditev potencialne prisotnosti protiteles proti virusu SARS-CoV-2. S ciljno izbranimi parametri (0,5 g/l Au, 5 l/min N2, 2,5 l/min H2) smo z USP sintetizirali AuND iz prekurzorja na osnovi zlatovega (III) klorida. AuND smo zbirali v steklenicah z demineralizirano vodo. Določili smo optimalne stabilizatorje in njihove koncentracije: 2,5 g/L PVP 40K ((C6H9NO)n) ter naknadno dodani 1 g/L trinatrijevega citrata (Na3C6H5O7) in saharoze (C12H22O11), da bi preprečili AuND aglomeracijo. Presevna elektronska mikroskopija AuND je pokazala, da so okrogli, s povprečno velikostjo okoli 50 nm in ozko velikostno porazdelitvijo, brez defektov in nizkim ZETA potencialom, 2,4 mV, pri pH 7,5. Za odstranitev vode iz suspenzije z AuND in stabilizatorji je bila uporabljena liofiliazcija, ki je bila izvedena pri naslednjih pogojih: zamrzovanje pri –40 °C, 4 h, atmosferski tlakprimarno sušenje pri +20 °C, 12 h, 1–4 Pasekundarno sušenje pri +30 °C, 33 h, 1–4 Pa. Nastali posušeni AuND so ohranili vse lastnosti, ki so potrebne za funkcijo oznak v hitrih LFIA testih: visoka površinska energija, površinska plazmonska resonanca in proste karboksilne skupine za konjugacijo. V nadaljevanju izvajanja eksperimentov smo, da bi potrdili doktorske hipoteze, izvedli raziskavo in optimizacijo konjugacije AuND z antigenom SARS-CoV-2, ki je bodičasti protein S1 (ak 1-681), in s protitelesi na zajčji IgG (RMG03). Za določitev poteka konjugacije AuND smo uporabili posredne (gelske elektroforeze, UV-Vis, preizkus delovanja) in direktne (XPS, SIMS) metode. Rezultati so pokazali, da se med našim postopkom konjugacije, v fazah čiščenja odstranimo večinski delež PVP in saharoze, ter s tem sprostimo proste karboksilne funkcionalne skupine citratnih ionov, ki so vezani na povšini AuND. Na teh prostih mestih lahko poteče konjugacija prek prostih -NH2 skupin na proteinih. Njihovo funkcionalnost smo ovrednotili z eksperimentalnimi testiranji lastno razvitega LFIA testa. Na osnovi tega smo postavili model in mehanizem konjugacije AuND, ki predpostavlja, da poteka konjugacija z antigeni oziroma protitelesi prek tvorbe peptidnih vezi med N-koncem proteina in prostimi karboksilnimi skupinami citratnega iona, ki je vezan na površino AuND prek svojih karboksilnih skupin. Za dodatno potrditev funkcionalizacije AuND z izbranimi biomolekulami smo v sklepnem delu doktorskega dela izvedli klinično študijo na pacientih UKC MB, ki je bila odobrena s sklepom KME RS št. 0120-148/2021/3. Rezultati študije so pokazali, da testi dosegajo visoko občutljivost (83 %) tako na vzorcih nosnega sluza kot na vzorcih seruma, slabšo specifičnost (66 % sluz, 74 % serum), visoko negativno napovedno vrednost (94 %) in slabšo pozitivno napovedno vrednost (38 % sluz, 45 % serum). S študijo smo potrdili delovanje AuND v funkciji oznak, saj so se testne in kontrolne linije v lastno razvitem LFIA hitrem testu obarvale.This doctoral thesis presents the studies, experimental work, characterisation techniques, results and analyses with the help of which we confirmed the hypothesis that it is possible to conjugate Au nanoparticles (AuNPs) prepared by the ultrasonic spray pyrolysis (USP) method with the target antigens of SARS-CoV-2 and recombinant IgG to function in a rapid lateral flow immunoserological assay (LFIA) to confirm the potential presence of antibodies against the SARS-CoV-2 virus. AuNPs were synthesised with the chosen parameters (0.5 g/l Au, 5 l/min N2, 2.5 l/min H2) from a precursor based on Au-chloride, using USP. AuNPs were collected in gas wash bottles filled with demineralized water. We determined the optimal stabilisers and their concentrations with the goal of preventing AuNPs agglomeration: 2.5 g/L PVP 40K ((C6H9NO)n), and subsequently added 1 g/L of trisodium citrate (Na3C6H5O7) and sucrose (C12H22O11). Microstructural analysis of the AuNPs showed them to be round, with sizes of around 50 nm and a narrow size distribution, without defects and with a low ZETA potential of 2.4 mV at a pH of 7.5. To remove water from the suspension with AuNPs and stabilisers, freeze-drying was used, which was carried out under the following conditions: freezing at -40 °C, 4 h, atmospheric pressureprimary drying at +20 °C, 12 h, 1-4 Pa, secondary drying at +30 °C, 33 h, 1-4 Pa. The resulting dried AuNPs retained all the properties required for their use as labels in LFIA assays: high surface energy, surface plasmon resonance, and free carboxyl groups for conjugation. To confirm the doctoral hypothesis, we continued with the experiments and the optimisation of the conjugation process of AuNPs with the SARS-CoV-2 S1 antigen, with the S1 (ak 1-681) subunit, and with anti-rabbit IgG antibodies (RMG03). Indirect methods such as gel electrophoresis, UV-Vis and LFIA experiments, and direct methods such as XPS and SIMS, were used to determine the mechanism of AuNPs conjugation. The results showed that during our conjugation process, the majority of PVP and sucrose is removed in the purification stages, thereby releasing free carboxyl functional groups of citrate ions, which are bound on the AuNPs surface. Conjugation happens via the free -NH2 groups of proteins and the vacant carboxyl sites on the surface of AuNPs. We evaluated their functionality by testing through our own LFIA test. Based on this, we set up a model and mechanism of AuNPs conjugation, which assumes that conjugation with antigens or antibodies takes place through the formation of peptide bonds between the N terminus of the protein and the free carboxyl groups of the citrate ion, which is bound to the AuNPs surface via its carboxyl groups. To further confirm the functionalisation of AuNPs with the selected biomolecules, in the concluding part of the doctoral thesis, we conducted a clinical study on patients of the University Medical Centre in Maribor, which was approved by The Commission of the Republic of Slovenia for Medical Ethics, with decision no. 0120-148/2021/3. The results of the study showed that the tests achieve a high sensitivity of 83% on both nasal mucus and serum samples and lower specificities of 66% (mucus) and 74% (serum), as well as a high negative predictive value of 94% and lower positive predictive values of 38% (mucus) and 45% (serum). Through the study, we confirmed the function of AuNPs as labels, with a successful staining of the test and control lines in our self-developed LFIA test

Simulation of a formation of the production line in reference to changing of the production orders

V diplomski nalogi so popisane osnove načrtovanja in planiranja proizvodnih linij in celic, ki so zelo odvisne od prostorske razmestitve in števila delovnih mest ter oblikovanega toka materiala. Za analizo izbrane proizvodne celice ter v nadaljevanju za izboljšavo te celice smo si pomagali s programom FlexSim. Glavni cilj naloge je ugotoviti, katera rešitev celice je najprimernejša glede na minimalne stroške izdelave enote izdelka. Za izboljšanje izbrane celice smo podali rešitve, ki so predstavljene v tretjem poglavju in jih nato pri izboljšavi tudi upoštevali. Ob upoštevanju rešitev smo naredili posodobljen simulacijski model celice in ga analizirali. Zaključek naloge predstavlja rezultate in ugotovitve izboljšane proizvodne celice.In this degree are described basics of designing and planning of the production lines and cells, that are very dependent of the spatial distribution, number of workplaces and formed material flow. For the analysis of the chosen production cell and in continuation for the improvement of this cell, the program FlexSlim was used as help. Main goal of this degree was to find out, which solution of the cell would be the most appropriate one, regarding the minimal costs of production of the product unit. For the improvement of the chosen cell were given solutions, which are presented in chapter three. These were also taken into account by the improvement. Considering these solutions, the modernized simulation model of the cell was made and analyzed. The final part of this degree shows the results and findings of the improved production cell

DEVELOPMENT OF WORKFLOWS ON THE GENERAL ADMINISTRATIVE PROCEDURE EXAMPLE

Ideja pisarne prihodnosti je že od nekdaj imeti na mizi samo terminal in tipkovnico. To pomeni, da mizo ne pokrivajo ogromne količine papirja. Glede na današnje zmogljivosti računalnikov in nizko ceno spominskega prostora, bi vso papirno dokumentacijo lahko spravili v digitalno obliko. S tem bi lahko prihranili ogromno denarja ter, poskrbeli za varnost in hranljivost dokumentov na enem samem mestu. Digitalizacija dokumentacije bi največ prednosti prinesla javnim ustanovam, saj njihov delovni proces vključuje ogromno raznih pogodb, odločb, prošenj, listin, zahtevkov, pritožb in podobnega. Izpolnjevanje, sledenje, organiziranje, arhiviranje in potrjevanje te dokumentacije je lahko zelo težavno in zamudno opravilo. Ideja diplomskega dela je, da iz poslovanja umaknemo papir in vse počnemo elektronsko, potrebne zadolžitve pri izpolnjevanju pa določimo v informacijsko podprtem poslovnem procesu ali delovnem toku posameznemu udeležencu v procesu, da izpolni določen del svoje naloge. Kot rezultat na koncu pričakujemo, da udeleženci delovnega procesa pri svojem delu ne bi uporabljali papirja. Posledično pa tudi večjo preglednost poslovnega procesa oziroma delovnega toka.The idea of an office of the future has always been about only having a computer and keyboard on your desk. This means it should not be covered in paper. Due to today’s computer efficiency and low prices of memory mediums, it is possible to transform paper documentation into a digital form. A lot of money could be saved with that, but at the same time, there is much better safety and everything can be stored conveniently in only one location. Public institutions would highly benefit from the digitalization of documents, because their workflow includes dealing with different contracts, provisions, applications, documents, claims, complaints etc on a daily basis. Completing, tracking, organizing and approving documentation is often a long and demanding process. The idea of this Thesis is to reduce the use of paper and work only electronically, appointing different tasks to individuals through information supported business process or workflow. Every individual present in the business process has to complete a task that is given to him. As a result we expect them not to use paper in their daily operations and also to have a much better transparency of their business process and working space

Informatization proposal of clear coatings manufacturing management process

Helios is a water-based coatings production company, located in Domžale. This thesis is related to Helios’s production unit in terms of optimizing the production process of the plant by applying means of informatization to several procedures. The company’s management sets the list of requirements to be satisfied, such as sub goals to be included in the solution proposed, or present management execution system information that will support the plant’s manufacturing process. Firstly, group Helios and company Helios, TBLUS are presented along with their manufacturing facility and IT support requirements. Further on, the theoretical and scientific concepts on which the practical part relies on will be described. In this part, planning, controlling and managing of production systems and processes are defined, mainly focusing on the IT support and management systems and enterprise modeling. The practical part of the thesis consists of two parts: (1) an exact replica of current business processes in the plant, including the description of working documentation and work places and (2) a suggested model of informatizing the manufacturing process, a procedure that represents the core of this thesis. In the proposal part the three-layered enterprise architecture model is analyzed including: business, application and technological layer. The application layer is described in detail, being the basis for implementation, functionalities and application modules. Concluding, it is argued that the solution proposed meets all expectations set and therefore fulfills the main objective and targets. Finally, in a closing section will be displayed advantages, potential vulnerabilities and risks that can occur throughout the implementation procedure of the proposed information system

Dropshipping method in an online store

V diplomskem delu Metoda dropshipping v spletni trgovini, smo najprej opisali teorijo spletne trgovine in to podkrepili s primeri različnih vrst le teh. V nadaljevanju smo opisali postopek izdelave spletne trgovine in s pomočjo teorije ter vodiča, v katerem smo to po korakih prikazali na spletni platformi Shopify. Na koncu smo razložili, kako poiskati donosno nišo za vaše lastno podjetje in predstavili postopek izbire pravega dobavitelja, kateri vam bo pomagal pri uresničevanju vaših načrtov.In the diploma with the title Dropshipping Method in an Online Store, the theory of an online store was described. There were also used different and variety of different types of an online store. In addition, we also described the process of creating an online store by using help of various theories, and step by step guide as is shown on Shopify online platform. In the conclusion of this diploma, we explained how to find a profitable niche for your own company, and presented the process of choosing the right supplier that will help you realizing your plans for the benefit of the company

OLAP ANALYSIS IN THE SCHOOL INFORMATION SYSTEM

Vsak dan se srečujemo z ogromno količino podatkov. Samo shranjevanje podatkov pa za uporabnike ne predstavlja nobene dodane vrednosti. Te podatke je potrebno analizirati in spraviti v smiselna poročila. Tematika diplomske naloge je uvajanje OLAP tehnologije v šolski informacijski sistem. Ravno ta tehnologija pa nam omogoča analizirati podatke in iz njih pridobiti dodatno vrednost. S tem bomo predstavili sam postopek modeliranja podatkovnega skladišča in postavitev tega na strežniku. Na primerih bomo predstavili ETL procese in težave, ki nastanejo pri teh transformacijah. Predstavili bomo tudi gradnjo OLAP kocke in njeno uporabo v MS Excelu. Cilj diplomske naloge je na resničnem projektu predstaviti proces uvajanja OLAP-a.We face a huge amount of data every day. Only storing that data does not create any added value for the users. It is necessary to analyze this data and convert it into meaningful reports. The theme of this diploma work is the introduction of OLAP technology in to a schools information system. This technology enables us to analyze the data and gain additional value from it. Using this procedure we will demonstrate the process of data warehouse modeling and the installation of said warehouse on a server. We will use examples to show ETL processes and problems that may arise with these transformations. We will also show the construction of a OLAP cube and its use with MS Excel. Our goal with this diploma work is to demonstrate the process of OLAP introduction on a real project

Developing Mobile Games with AndEngine

V diplomskem delu je predstavljen razvoj igre s pomočjo knjižnice AndEngine. Knjižnice vsebujejo različne funkcionalnosti, ki jih prikličemo oz. uporabimo pri samem razvoju. Ni nam treba vedeti, kako točno funkcionalnost deluje, ampak moramo vedeti samo, kako funkcionalnost priklicati oz. uporabiti. Zato je uporaba knjižnic za razvijalce priporočljiva, saj s tem dosežemo večjo stabilnost in zanesljivost igre, prav tako pa nam olajšajo delo. Preučili bomo knjižnico AndEngine, v kateri bomo spoznali življenjski cikel, ustvarjanje različnih aktivnosti, delo z entitetami, modifikatorje, kamere in dodatne razširitve za samo knjižnico. Predstavili in analizirali bomo konkurenčne knjižnice in naredili preprost prototip igre, ki bo namenjena otrokom za učenje ločevanja odpadkov.This diploma paper presents game development using the AndEngine library. Libraries include different functions, which we launch and use during the development. We do not have to be familiar with the functions in particular, we simply have to know how to launch and use the function. Therefore, the use of libraries is recommended for developers as we achieve greater stability and reliability of the game, and it also facilitates our work. We will study the AndEngine library, in which we will get to know the life cycle, activity creating, working with entities, modifiers, cameras and different library extensions. We will present and analyze competitive libraries, and develop a simple game prototype intended to teach children waste sorting

Study of gold nanoparticles conjugated with SARS-CoV-2 S1 spike protein fragments

This study reports on the successful conjugation of SARS-CoV-2 S1 spike protein fragments with gold nanoparticles (AuNPs) that were synthesised with Ultrasonic Spray Pyrolysis (USP). This method enables the continuous synthesis of AuNPs with a high degree of purity, round shapes, and the formation of a surface that allows various modifications. The conjugation mechanism of USP synthesized AuNPs with SARS-CoV-2 S1 spike protein fragments was investigated. A gel electrophoresis experiment confirmed the successful conjugation of AuNPs with SARS-CoV-2 S1 fragments indirectly. X-ray Photoelectron Spectroscopy (XPS) analysis confirmed the presence of characteristic O1s and N1s peaks, which indicated that specific binding between AuNPs and SARS-CoV-2 S1 spike protein fragments takes place via a peptide bond formed with the citrate stabiliser. This bond is coordinated to the AuNP’s surface and the N-terminals of the protein, with the conjugate displaying the expected response within a prototype LFIA test. This study will help in better understanding the behaviour of AuNPs synthesised with USP and their potential use as sensors in colorimetric or electrochemical sensors and LFIA tests