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Two-Dimensional Microfluidic Bioarray for Nucleic Acid Analysis
Published in Iniewski Krzysztof, Integrated Microsystems, 2017
Nucleic acid hybridization techniques feature the use of a probe nucleic acid molecule to detect a target nucleic acid molecule. Here, probe molecules are usually short single-stranded nucleic acids (DNA or RNA) or oligonucleotides with known sequences; whereas target molecules are prepared from polymerase chain reaction (PCR) amplification of genomic extracts. Probe-target hybridization leads to the formation of a double-stranded molecule, called duplex. The concept of DNA microarray was evolved from Southern blotting technology based on solid-phase hybridization in the early 1990s [1]. This method relies on the immobilization of the probe molecules onto the solid surface to recognize their complementary DNA target sequence by hybridization. Up to millions of features have been integrated into a standard glass slide or silicon chip by microprinting or in situ synthesis of oligonucleotides [2,3]. The relative abundance of nucleic acid sequences in the target can be measured from chip-hybridization results optically, electrochemically, or radioactively, with proper detection labels [4]. DNA microarrays have dramatically accelerated many types of investigations including gene expression profiling, comparative genomic hybridization, protein–DNA interaction study (chromatin immunoprecipitation), single-nucleotide polymorphism (SNP) detection, and nucleic acid diagnostic applications. The advances in DNA microarray technology during the last couple of years have been summarized in many books and reviews [4–8].
In situ Hybridization Histochemistry
Published in Edythe D. London, Imaging Drug Action in the Brain, 2017
Martin K.-H. Schofer, James P. Herman, Stanley J. Watson
The theoretical framework around which the in situ hybridization method is built is borrowed extensively from the dissociation-reassociation kinetics of DNA and RNA in solution. (The kinetics of nucleic acid hybridization are discussed in detail elsewhere: Britten and Davidson, 1985; Wetmur and Davison, 1968). For the most part, hybridization in situ, using in essence mRNA immobilized in tissue sections, is assumed to obey kinetics similar to hybridization in solution. While there are several reasons to believe this may not be entirely true (such as uncertainty as to the accessibility of cellular mRNA for hybridization in situ; see below), in general, this assumption has provided a good working model for design and analysis of the in situ hybridization method.
Luminescent Lanthanide Probes as Diagnostic and Therapeutic Tools
Published in Astrid Sigel, Helmut Sigel, Metal Ions in Biological Systems, 2004
Advances in recombinant DNA technology and sensitive methods for analyzing the organization of specific genes are major contributors to genomics. These techniques rely on nucleic acid hybridization probes for the detection of complementary nucleic acid sequences. DNA probes are more and more used in diagnostic medicine, e.g., in the recognition of genetic predisposition to a given disease, virus detection, bacterial identification or antibiotic sensitivity testing. Until the 1990’s, radioisotopes (e.g., 32P, 3H) were common labels for nucleic acid probes, but stability and safety problems led to search for alternative, non-isotopic stains. Truly biochemical methods were proposed, such as biotinylated nucleotide analogues detected by streptavidin or proteins crosslinked to the nucleic acid, but their sensitivity never matched the sensitivity of radio-assays, leaving an opportunity for luminescent assays.
An insight into clinical and laboratory detections for screening and diagnosis of cervical cancer
Published in Expert Review of Molecular Diagnostics, 2023
Shruthi Padavu, Pooja Aichpure, Ballamoole Krishna Kumar, Anoop Kumar, RadhaKanta Ratho, Shipra Sonkusare, Indrani Karunasagar, Iddya Karunasagar, Praveen Rai
The nucleic acid-based detection of HPV has resulted in a significant revolution in the discovery of novel HPV types, epidemiological research, facilitation of diagnosis in the healthcare system, and monitoring immunization programs. The main objective of these assays is to identify HPV DNA from cervical samples due to their high sensitivity and specificity [38]. The error-free identification and typing of HPVs are essential for accurately predicting the premalignant state of cervical cancer. PCR, qPCR, and isothermal nucleic acid amplification tests are some of the most frequently used methods for detecting HPV DNA as an etiological agent of cervical cancer [39]. Based on their principles, these molecular detection tests can be classified into two categories: a) nucleic acid hybridization tests, and b) nucleic acid amplification tests. Many diagnostic kits approved by the Food and Drug Administration (FDA) based on nucleic acid detection techniques are commercially available. Table 2 lists the principles and applications of different such kits to detect HR-HPV.
Molecular detections of coronavirus: current and emerging methodologies
Published in Expert Review of Anti-infective Therapy, 2022
Mingkun Diao, Lang Lang, Juan Feng, Rongsong Li
Biosensors are devices with transducers that can convert biological response into electrical, electrochemical or optical signals [35,36]. Unlike above described methods, biosensor detection of RNA are based on nucleic acid hybridization rather than amplification [37]. When viral RNA sequences hybridize with a probe (synthetic single-stranded DNA), the biochemical signals produced by the formation of DNA–RNA complex are captured and converted into detectable electrical, electrochemical or optical signals to report the presence of specific RNA sequence. Commonly used electrical and electrochemical biosensors in viral detections include cyclic voltammetric (CV) [38], square-wave voltammetry (SWV) [39], chronoamperometry (CA) [40], electrochemical impedance spectroscopy (EIS) [41], and differential pulse voltammetry (DPV) [42]; while surface plasmon resonance (SPR) [43], fluorescence [44], silicon microring resonator (SMR) [45], and colorimetry [46] are frequently used readout in optical biosensors.
Molecular techniques for the genomic viral RNA detection of West Nile, Dengue, Zika and Chikungunya arboviruses: a narrative review
Published in Expert Review of Molecular Diagnostics, 2021
Antonio Mori, Elena Pomari, Michela Deiana, Francesca Perandin, Sara Caldrer, Fabio Formenti, Manuela Mistretta, Pierantonio Orza, Andrea Ragusa, Chiara Piubelli
The direct molecular methods to detect viral genomes can be roughly summarized in: i) Target Analysis (TA), including the nucleic acid amplification (T-NAA) and the no-amplified nucleic acid hybridization probes (T-nAHP) techniques; ii) no Target analysis (nTA) such as the Next-Generation Sequencing (NGS) (Figure 2). The NAA techniques have a better analytical sensitivity in comparison to NGS, and the target amplification techniques commonly employed are based on polymerase chain reaction (PCR) and isothermal methods. However, great strides have been made in the development and improvement of hybridization techniques to achieve a very efficient and sensitive high-throughput systems, such as DNA arrays (micro- and macro-arrays). (Tables 1 and 2) collect different molecular methods described in literature for arboviruses detection and discussed in the following paragraphs.