br Results br Discussion br Materials

Results
Discussion
Materials and Methods
Introduction Deoxyribonucleic BRD4770 receptor (DNA) is among the most important biopolymers in living organisms alongside carbohydrates and proteins. DNA is a linear polymer consisting of four types of nucleobases, deoxyribose, and phosphate. The backbone of DNA is made up of alternating deoxyribose and phosphate while the four nucleobases, i.e. adenine (A), thymine (T), cytosine (C), and guanine (G), form the reactive sites for DNA to take part in encoding, transmitting, and expressing genetic information. Two strands of DNA run in opposite directions with the four nucleobases pairing up with each other via hydrogen bonds according to Watson-Crick pairing rules to give DNA its double helix structure [1]. In living organisms, genetic information is conveyed through the order of the four nucleobases. In other words, the arrangements of the nucleobases along the backbone of DNA in specific orders are the mechanism of preserving and transmitting genetic information. Located in nucleus and mitochondria [2], DNA is able to unwind its strands where single-stranded DNA (ss-DNA) serves as a template for transcription, thereafter allowing the newly synthesized strand to undergo translation modifications before serving its function in cells. Despite its high fidelity rate, DNA can be damaged because it is under constant attack both endogenously and exogenously, creating around ten thousand lesions in its structure in a single cell per day [3]. There are many types of DNA lesions. Ranging from minor to major modifications in its chemical structure, these lesions occur through the formation of abasic sites, base mismatch, and the breakage of single- or double-stranded DNA (ds-DNA) (Fig. 1) [4]. Not only do they cause genetic information to be lost, but also can change the genome irreversibly as permanent mutations [5]. Consequently, mutagenesis and/or carcinogenesis occur and consequently induce hereditary disorders, cancer, and non-heritable somatic cell diseases after surpassing certain thresholds [6], [7].
Sources of DNA lesions and their repair DNA lesions can be classified into endogenous and exogenous types. Compared to exogenous lesions, endogenous lesions are more frequent. Endogenous lesions are caused by a variety of factors such as spontaneous or enzymatic conversions, reactive oxygen species (ROS), and alkylating agents [8]. Exogenous lesions are due to exposures to environmental factors like toxins and mutagenic compounds, ultraviolet (UV) light, radioactive materials, and relatively high temperature. Each factor will be discussed briefly to aid the understanding of the extent of DNA damage.
DNA glycosylase assays Towards the ultimate goal of improving human health, the activity of DNA glycosylases needs to be closely monitored to gain a deep understanding of the restoration process of damaged DNA and to aid the design of therapeutic strategies. In recent years, intensive research efforts have been directed to the development of DNA glycosylase assays hoping to circumvent the limitations of conventional DNA glycosylase assays like gel electrophoresis. Among all DNA glycosylase assays reported, colorimetric, fluorometric, and electrochemical DNA glycosylase assays are most intensively investigated. Colorimetric assays are based on visual inspection of color changes or spectrometric measurement of absorption. They have the advantages of straightforwardness, low cost, and sometimes instrument-free (for example pregnancy test strips), but their low sensitivity is a major obstacle in the development of DNA glycosylase assays. Fluorometric assays rely on the measurement of light emitted by fluorophores (fluorescent molecules or quantum dots). When light of appropriate wavelengths illuminates on the fluorophores, it is absorbed by the fluorophores, thereby promoting electrons of the fluorophores from their ground states to excited states. Upon relaxing to the ground states, light of longer wavelengths is emitted in the process. Fluorometric assays have the advantage of good sensitivity, but the complicated optics of fluorometers often limits their use in centralized laboratory. Chemiluminescent assays leverage on chemical reactions that produce intermediates in their electronically excited states. Light is emitted when those intermediates eventually return to their ground states [42]. Apart from their high sensitivity, unlike fluorometric or colorimetric assays where a light source and complicated optics are required, the main advantages of chemiluminescent assays are their simplicity of instrumentation and high sensitivity. Unfortunately, there are not many chemiluminescent compounds available, and hence chemiluminescence is only limited to a relatively small number of assays. On the other hand, to widen the applications of DNA glycosylase assays and eventually reach point-of-care, it is vital that the developed DNA glycosylase assays are simple, fast, highly selective and sensitive, affordable, and portable. In this regard, electrochemical assays are capable of meeting the above-mentioned criteria because of their inherently high portability, high sensitivity, and excellent compatibility with microfabrication technology. A good example of electrochemical assays is blood sugar assay. Before personal glucose meters reached the market, blood sugar tests had to be conducted in clinical laboratories.