Chemistry / Kimya
Permanent URI for this collectionhttps://hdl.handle.net/11147/4072
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Article Free-Standing Three-Dimensional Graphene Scaffolds for Protease Functional Assay(Elsevier Science Sa, 2024) Ng, Zhi Kai; Yan, Evelias; Goyal, Garima; Gudlur, Sushanth; Kanagavel, Deepankumar; Yildiz, Umit Hakan; Teo, Edwin Hang TongThree-dimensional graphene scaffolds (3d-GS) of high porosity possessing good fluorescence quenching properties are potential candidates for the development of optical biosensors. Herein, we demonstrate the feasibility of utilising intact and free-standing 3d-GS for sensitive detection of proteases, a class of disease diagnosis bio-markers of significant interest. Recombinant OmpT was employed as a model protease for validating the pro-posed methodology. A short (15-residue) peptide sequence encoding a specific recognition site for OmpT was end-labelled with a fluorescent dye (5-FAM) whose fluorescence is quenched when the peptide is anchored to 3d-GS. However, in the presence of OmpT, the peptide is cleaved and released from 3d-GS, resulting in a sig-nificant recovery in fluorescence. The functional assay described herein involves a single step fabrication process of anchoring the peptide to 3d-GS. The integrity of the 3d-GS is hypothesised to overcome the concern of dynamic requenching associated with the typical homogeneous assays based on graphene, yielding a limit of detection (LOD) of similar to 140 nM, which is over an order higher than homogeneous assays performed using the same composition of graphene in powdered form. To the best of our knowledge, this is the first report on utilising free-standing 3d-GS for facile assaying of proteases.Article Citation - WoS: 4Citation - Scopus: 4Colorimetric Assaying of Exosomal Metabolic Biomarkers(MDPI, 2023) Yan, Evelias; Goyal, Garima; Yıldız, Ümit Hakan; Boehm, Bernhard O.; Palaniappan, AlagappanExosomes released into the extracellular matrix have been reported to contain metabolic biomarkers of various diseases. These intraluminal vesicles are typically found in blood, urine, saliva, breast milk, cerebrospinal fluid, semen, amniotic fluid, and ascites. Analysis of exosomal content with specific profiles of DNA, microRNA, proteins, and lipids can mirror their cellular origin and physiological state. Therefore, exosomal cargos may reflect the physiological processes at cellular level and can potentially be used as biomarkers. Herein, we report an optical detection method for assaying exosomal biomarkers that supersedes the state-of-the-art time consuming and laborious assays such as ELISA and NTA. The proposed assay monitors the changes in optical properties of poly(3-(4-methyl-3'-thienyloxy) propyltriethylammonium bromide) upon interacting with aptamers/peptide nucleic acids in the presence or absence of target biomarkers. As a proof of concept, this study demonstrates facile assaying of microRNA, DNA, and advanced glycation end products in exosomes isolated from human plasma with detection levels of ~1.2, 0.04, and 0.35 fM/exosome, respectively. Thus, the obtained results illustrate that the proposed methodology is applicable for rapid and facile detection of generic exosomal biomarkers for facilitating diseases diagnosis.