A self-assembling split aptamer multiplex assay for SARS-COVID19 and miniaturization of a malachite green DNA-based aptamer

O’Steen, Martin R.; Kolpashchikov, Dmitry M.

doi:10.1016/j.snr.2022.100125

Cited by 3 publications

(2 citation statements)

References 43 publications

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“…Light-up aptamers have been previously combined in different nucleic acid circuits for specific diagnostics and detection including for single nucleotide variants, pathogenic bacteria, and SARS-CoV-2 RNA. − Generally, these methods rely on direct binding of the aptamer to a small amount of the target or require prior amplification of the target before detection. , Both strategies lack the real-time quantification capabilities observed for qPCR. Our method combines the qPCR advantages with light-up dye and aptamer-based detection.…”

Section: Resultsmentioning

confidence: 99%

“… 28 − 31 Generally, these methods rely on direct binding of the aptamer to a small amount of the target 28 or require prior amplification of the target before detection. 31 , 32 Both strategies lack the real-time quantification capabilities observed for qPCR. Our method combines the qPCR advantages with light-up dye and aptamer-based detection.…”

Section: Resultsmentioning

confidence: 99%

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DNA-Aptamer-Based qPCR Using Light-Up Dyes for the Detection of Nucleic Acids

Goyal,

Singh,

Sengupta

et al. 2023

ACS Omega

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Quantitative polymerase chain reaction (qPCR) is widely used in detection of nucleic acids, but existing methods either lack sequence-specific detection or are costly because they use chemically modified DNA probes. In this work, we apply a DNA aptamer and light-up dye-based chemistry for qPCR for nucleic acid quantification. In contrast to the conventional qPCR, in our method, we observe an exponential decrease in fluorescence upon DNA amplification. The qPCR method we developed produced consistent Ct vs log 10 (DNA amount) standard curves, which have a linearfit with R 2 value > 0.99. This qPCR technique was validated by quantifying gene targets from Streptococcus zooepidemicus ( SzhasB ) and Mycobacterium tuberculosis ( MtrpoB ). We show that our strategy is able to successfully detect DNA at as low as 800 copies/μL. To the best of our knowledge, this is the first study demonstrating the application of light-up dyes and DNA aptamers in qPCR.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

DNA-Aptamer-Based qPCR Using Light-Up Dyes for the Detection of Nucleic Acids

Goyal,

Singh,

Sengupta

et al. 2023

ACS Omega

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Effect of ionic strength on DNA–dye interactions of Victoria blue B and methylene green using UV–visible spectroscopy

Rahman,

Khan,

Rahman

et al. 2023

Zeitschrift Für Physikalische Chemie

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Victoria blue and methylene green dyes have both been extensively studied due to their numerous applications, including their ability to bind to DNA. Dyes are very important in everyday life with applications in textile, cosmetics, food and pharmaceutical industries. It has been found that some of them adversely affect human health causing severe abnormalities. Among these abnormalities, cancer is of great concern due to its fatal and almost non-recoverable nature. In this work we have studied the binding of two dyes namely Victoria blue B (VBB) and Methylene green (MG) with double stranded DNA (Salmon sperm). The interactions were studied in the presence of different concentrations of buffer solutions at a constant pH. The selected dyes showed interactions with double-stranded DNA through intercalation and electrostatic modes. Upon increasing ionic strength of the buffer the binding constant (K b ) value for MG was decreased whereas increased for VBB, which conclude that, at higher ionic strength (0.5 M) the DNA–MG interactions is lower and DNA–VVB interactions is maximum. The carcinogenicity of a given dye is indicated from its binding constants in the current study. Based on the recorded K b values of the selected dyes it was concluded that proper disposing and precautions should be taken while utilizing/dealing these dyes in order to minimize/avoid their impact on environment and human health.

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A DAMP-Based Assay for Rapid and Affordable Diagnosis of Bacterial Meningitis Agents: Haemophilus influenzae, Neisseria meningitidis, and Streptococcus pneumoniae

Shkodenko,

Mohamed,

Ateiah

et al. 2024

IJMS

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The rapid and accurate diagnosis of meningitis is critical for preventing severe complications and fatalities. This study addresses the need for accessible diagnostics in the absence of specialized equipment by developing a novel diagnostic assay. The assay utilizes dual-priming isothermal amplification (DAMP) with unique internal primers to significantly reduce non-specificity. For fluorescence detection, the dye was selected among Brilliant Green, Thioflavin T, and dsGreen. Brilliant Green is preferred for this assay due to its availability, high fluorescence level, and optimal sample-to-background (S/B) ratio. The assay was developed for the detection of the primary causative agents of meningitis (Haemophilus influenzae, Neisseria meningitidis, and Streptococcus pneumoniae), and tested on clinical samples. The developed method demonstrated high specificity, no false positives, sensitivity comparable to that of loop-mediated isothermal amplification (LAMP), and a high S/B ratio. This versatile assay can be utilized as a standalone test or an integrated assay into point-of-care systems for rapid and reliable pathogen detection.

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A self-assembling split aptamer multiplex assay for SARS-COVID19 and miniaturization of a malachite green DNA-based aptamer

Cited by 3 publications

References 43 publications

DNA-Aptamer-Based qPCR Using Light-Up Dyes for the Detection of Nucleic Acids

DNA-Aptamer-Based qPCR Using Light-Up Dyes for the Detection of Nucleic Acids

Effect of ionic strength on DNA–dye interactions of Victoria blue B and methylene green using UV–visible spectroscopy

A DAMP-Based Assay for Rapid and Affordable Diagnosis of Bacterial Meningitis Agents: Haemophilus influenzae, Neisseria meningitidis, and Streptococcus pneumoniae

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