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dc.contributor.authorAUGUSTINE, SHINE-
dc.date.accessioned2022-02-21T08:25:53Z-
dc.date.available2022-02-21T08:25:53Z-
dc.date.issued2020-08-
dc.identifier.urihttp://dspace.dtu.ac.in:8080/jspui/handle/repository/18791-
dc.description.abstractNanomaterials can be defined as the materials whose dimensions fall in the range of 1-100 nm, exhibiting peculiar chemical, physical and molecular properties compared to the bulk counterpart. They have proven to be of significant interest in the field of physical, chemical and biological systems. Among the different nanomaterials, transition metal oxides (TMOs) have emerged as a new class of material towards the advancement of technology in different domains. TMOs are composed of oxygen atoms bound to the transition metals leading to different physiochemical characteristics and polymorphism. With respect to their bulk counterpart, the nanostructured transition metal oxides (nTMOs) have attracted enormous interest due to their high surface activity, better catalytic efficiency, remarkable electrochemical properties, and variable oxidation states. Among the various nTMOs, molybdenum trioxide (MoO 3 ) has gained immense attraction in the field of material science. The nanostructured MoO 3 possess outstanding physical, chemical and electronic properties such as good electrical conductivity, tuneable band gap, optical and high catalytic activity etc. Owing to these unique characteristics, MoO 3 has found many applications including electrochromic systems, energy storage units, superconductors, thermal materials, antibacterial, gas and biomedical sensors. Moreover the different oxidation states of MoO 3 ranging from +2 to +6 aids in the formation of different conducting oxide states that facilitate free movement of electrons and depending on the application, they can be functionalised using stable chemical linkers (such as APTES,MPTES and Serine etc.) Cancer is currently a serious concern and is a medical threat to the contemporary world. In 2018, around 8.1 million cases and 9.6 million cancer deaths were reported (GLOBOCAN) aggravating the need for its early diagnosis. As reported by the International Agency for Research on Cancer (IARC), the trend is likely to increase due to the limited 12 availability of diagnostic equipment. Among the major cancer, types (Prostate, Breast, Leukaemia, Colon, Lung, Liver etc.) breast cancer (BC) is the most frequent in women worldwide and may be curable in ~70–80% of patients in early-stages of the disease. It presently accounts for about 11.6 % of all the female cancers and 22.9% of invasive cancers in women. Several conventional techniques such as biopsy, immunohistopathology, Fluorescence in situ hybridization (FISH), Enzyme linked immunosorbent assay (ELISA) and mammography are currently utilised for breast cancer detection. These techniques are quite complex and also necessitate collection of a tissue sample by an experienced surgeon, analysis by physician and pathologist, requires at least 8-10 days for the analysis. Additionally, they are expensive and may yield false-negative result rates with a limited potential for early diagnosis of the disease. For the last two decades, simple and fast analytical techniques based on electrochemical sensors have been found to have considerable potential due to their high sensitivity, selectivity, lower detection limits, multi target analyses, reduced costs of testing, low power consumption, simple instrumentation and point-of-care testing. Moreover, the detection of clinical biomarkers plays a crucial role in the detection of breast cancer, design of individual therapies, and to identify underlying processes involved in the disease. Among the various known biomarkers for breast cancer, Estrogen receptor (ER), Progesterone receptor (PR) and Human epidermal growth factor receptor 2 (HER-2) are some of the relevant biomarkers for the diagnosis and routine monitoring of breast cancer. An individual who is prone to breast cancer undergoes this triple marker test. Moreover, it may be helpful in deciding the treatment strategies (Endocrine or Trastuzumab therapy) for the breast cancer patient. HER-2 is a transmembrane protein comprising of tyrosine kinase receptor of 185 kDa that is responsible for the growth and differentiation of the breast cells. The extracellular 13 domain of HER-2 is cleaved into serum, leading to the fluctuation in its concentration indicating the presence of the disease. The higher levels of HER-2 are associated with overexpression of HER-2 (20-30%), increased tumour burden resulting in the emergence of breast cancer. The average level of HER-2 in serum is determined to be around ~ 15 ng mL -1 and it rises to ~75 ng mL -1 in the advanced stage. Thus, monitoring of HER-2 levels in serum can be utilised for the timely diagnosis and prognosis of this disease. In the present thesis, attempts have been made to fabricate label-free, simple, biocompatible biosensing platforms based on nTMOs (nMoO 3 ) and its hybrid (MoO 3 @RGO and MoO 3 @NH 2 -MWCNTs) for the detection of breast cancer serum biomarker (HER-2). In this context, efforts have been made to synthesize nMoO 3 via one-pot hydrothermal method and subsequently functionalise with APTES linker molecules for covalent attachment of HER-2 biomolecules. Further, to improve the conductivity and electroactive surface area, these nMoO 3 was incorporated into a 2D carbon substrates (reduced graphene oxide, RGO) resulting in better charge transfer ability, increased mechanical stability, and efficient heterogeneous electron activity with the improved surface area. Likewise, to further enrich the biosensing parameters such as better sensitivity, detection limit and with reduced steps of fucntionalisation, amine functionalised multi-walled carbon nanotubes (-NH 2 -MWCNTs) was introduced along with the MoO 3. The aminated MWCNT aided in simpler fabrication strategy by reducing the requirement of linker needed for bioconjugation. This nanocomposite exhibited excellent electrochemical conductivity with enhanced heterogeneous electron activity towards HER-2 detection. The results obtained using the fabricated biosensors in the clinical samples (serum) are in good agreement with the ELISA results. 14 The thesis is divided into six Chapters as described below: Chapter 1 on “Introduction and Literature Review” highlights the importance of transitional metal oxides, their characteristic properties and application with emphasis on its utility as biosensors. Besides this, attempts have been made to give a detailed literature review on the different aspects of transitional metal oxides (nTMOs) and hybrids with carbonaceous materials (RGO, MWCNTs) and their functionalization with linker molecules. Efforts have been made to highlight the importance of breast cancer diagnosis along with the existing gaps in the current detection systems. Further, the salient features of antibodies used for fabrication of electrochemical immunosensor for the detection of breast cancer biomarker have also been discussed. Chapter 2 based on “Materials and Experimental Techniques” gives details of the various experimental techniques such as Fourier transform infra red (FTIR) spectroscopy, Raman spectroscopy, X-ray Diffraction (XRD), Brunauer-Emmett-Teller (BET), Scanning electron microscopy (SEM), Transmission electron microscopy (TEM), X-ray photoelectron spectroscopy (XPS) and Atomic force microscopy (AFM) that have been used to characterize the nanostructured molybdenum trioxide (nMoO 3 ) and its hybrids with RGO and NH 2 - MWCNTs based electrodes and immunoelectrode. Electrochemical techniques such as cyclic voltammetry (CV), differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS) have been utilized to characterize these nTMOs based electrodes and immunoelectrode. Electrochemical studies of the fabricated immunoelectrode as a function of HER-2 concentration have been conducted using DPV technique. Attempts have been made to describe the different procedures and protocols used to estimate the figure of merit of nTMOs based immunosensor for breast cancer biomarker (HER-2) detection. 15 Chapter III unravels the results of the studies relating to the fabrication of emerging nanostructured molybdenum trioxide (nMoO 3 ) based immunosensor. The synthesis of nMoO 3 using one pot hydrothermal method followed by its functionalization using APTES linker. The electrophoretically fabricated APTES/nMoO 3 /ITO electrode was covalently immobilized with anti-HER-2 using EDC-NHS linker chemistry. The results of SEM and TEM studies have revealed the one-dimensional (nanorods) morphology of the as-synthesised nanomaterial. The biocompatible behaviour was investigated using human HEK 293T cell lines. The results of electrochemical studies of the proposed immunosensor revealed high sensitivity (0.904 μA mL ng -1 cm -2 ), wide linear detection range (2.5-110 ng mL -1 ) and a lower detection limit of 2.47 ng mL -1 with a shelf life of 30 days. The response of immunosensor was validated using serum samples of breast cancer patient and were found to be in acceptable limit. This is the first report on the biosensing application of the synthesised nMoO 3 that exhibited a simple, label free and biocompatible biosensor. In order to further, improve the sensitivity and linear range of the fabricated immunosensor nMoO 3 was incorporated onto the 2D electroactive reduced graphene oxide (RGO) sheet to form a nanohybrid resulting in enhanced electrochemical properties. Chapter IV describes results of the studies relating to the in situ preparation of the MoO 3 prepared onto the reduced graphene oxide (RGO) via one-pot low-temperature hydrothermal synthesis and further functionalized via APTES. RGO is an electroactive material containing various active sites wherein the nucleation of MoO 3 starts and results in the growth of nanorods under hydrothermal conditions. The MoO 3 prevents restacking of the RGO sheets, providing room for enhanced electron mobility by shuttling mechanism. The surface area of this hybrid (258 m 2 g −1 ) was found to be 14 times greater than that of pristine nMoO 3 (17.19 m 2 g −1 ) and the pore size was enhanced by four times. Thin films of this nanohybrid (APTES/nMoO 3 @RGO) was deposited onto ITO coated glass substrate using 16 electrophoretic deposition (40 V for 2 min) technique. Subsequently, the monoclonal antibodies (anti-HER-2) were immobilised via EDC-NHS covalent chemistry onto the APTES/MoO 3 @RGO/ITO electrode. The APTES/MoO 3 @RGO/ITO electrode has shown improved heterogeneous electron transfer (>1.5 times) with respect to that of the APTES/ nMoO 3 /ITO electrode indicating faster electron transfer kinetics. This nanohybrid based immunosensor sensor BSA/anti-HER-2/APTES/MoO 3 @RGO/ITO has shown improved parameters i.e. higher diffusion coefficient; enhanced heterogeneous electron transfer and improved biomolecular loading resulting in broader linear detection range (0.001-500 ng mL −1 ), better sensitivity (13 µA mL ng −1 cm −2 ) and high selectivity. Moreover, the remarkable lower limit of detection 0.001 ng mL −1 revealed that this sensor could be used to detect even minute concentration of HER-2 biomolecules in the physiological range. Further efforts have been made to fabricate a simpler, reliable biosensor with enhanced sensitivity and improved shelf life towards efficient detection of breast cancer biomarker. For this purpose, amine functionalised multi-walled carbon nanotubes (NH 2 -MWCNT) was introduced along with the MoO 3 leading to the nanocomposite formation. Chapter V demonstrates the results of the studies relating to the fabrication of an electrochemical biosensor based on the as synthesised nanohybrid of MoO 3 @NH 2 -MWCNTs via one-pot hydrothermal synthesis. The NH 2 -MWCNTs is known to possess excellent electrical properties, high aspect ratio, large specific surface area and faster electron transport. Moreover, the incorporation of MoO 3 with NH 2 -MWCNTs results in superior immunosensing platform leading to improved electrochemical performance and better sensitivity towards the detection of HER-2. The SEM studies of the as synthesised nanocomposite reveal the formation of nanorods wrapped with thin fibres of –NH 2 -MWCNTs forming a dense network. In addition, the -NH 2- MWCNTs resulted in enhanced heterogeneous electron transfer (~10 times) compared to pristine MoO 3 with an average 17 surface area of 63 m 2 g -1 . This fabricated immunosensing platform using anti-HER-2 as antibody towards HER-2 detection exhibited remarkable sensitivity of about 26 µA mL ng −1 cm −2 per decade in the dynamic linear range (10 -6 –10 3 ng mL -1 ) and shelf life of about five weeks when stored at 4 °C. Thus, the MoO 3 @NH 2 -MWCNTs composite have shown excellent electrochemical behaviour (sensitivity and linear range) with respect to APTES/nMoO 3 and APTES/MoO 3 @RGO based electrodes and have potential to be utilised as immunosensing matrix for the detection of other cancer analytes including ovarian, lung etc. Chapter VI on “Summary and future prospects” highlights the role of nTMOs and its hybrids and their arrangement in the fabrication of efficient biosensors for breast cancer biomarker (HER-2) detection. Further, it also discusses the future prospects pertaining to the commercialization aspects for development of flexible platforms and towards multi analyte detection.en_US
dc.language.isoenen_US
dc.publisherDELHI TECHNOLOGICAL UNIVERSITYen_US
dc.relation.ispartofseriesTD - 5298;-
dc.subjectNANOMATERIALS BASED BIOSENSORSen_US
dc.subjectBREAST CANCER DETECTIONen_US
dc.subjectINTERNATIONAL AGENCY FOR RESEARCH ON CANCER (IARC)en_US
dc.subjectTRANSITION METAL OXIDES (TMOs)en_US
dc.titleDEVELOPMENT OF NANOMATERIALS BASED BIOSENSORS FOR BREST CANCER DETECTIONen_US
dc.typeThesisen_US
Appears in Collections:Ph.D. Bio Tech

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