Low-Cost Arduino Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) for Sensitive Nucleic Acid Detection.

This work presents a low-cost transcription loop-mediated isothermal amplification (RT-LAMP) instrument for nucleic acid detection, employing an Arduino Nano microcontroller. The cooling system includes customized printed circuit boards (PCBs) that serve as electrical resistors and incorporate fans. An aluminum block is designed to accommodate eight vials. The system also includes two PCB heaters-one for sample heating and the other for vial lid heating to prevent condensation. The color detection system comprises a TCS3200 color 8-sensor array coupled to one side of the aluminum heater body and a white 8-LED array coupled to the other side, controlled by two Multiplexer/Demultiplexer devices. LED light passes through the sample, reaching the color sensor and conveying color information crucial for detection. The top board is maintained at 110 ± 2 °C, while the bottom board is held at 65 ± 0.5 °C throughout the RT-LAMP assay. Validation tests successfully demonstrated the efficacy of the colorimetric RT-LAMP reactions using SARS-CoV-2 RNA amplification as a sample viability test, achieving 100% sensitivity and 97.3% specificity with 66 clinical samples. Our instrument offers a cost-effective (USD 100) solution with automated result interpretation and superior sensitivity compared to visual inspection. While the prototype was tested with SARS-CoV-2 RNA samples, its versatility extends to detecting other pathogens using alternative primers, showcasing its potential for broader applications in biosensing.

Substances injected at the Sydney supervised injecting facility: A chemical analysis of used injecting equipment and comparison with self-reported drug type.

Providing information about substances injected can reduce the negative impact of illicit drug consumption and support people who inject drugs to make informed decisions. In Australia, information about drugs injected relies largely on periodic self-report surveys. For the first time, the analysis of the residual content of used injecting equipment was conducted in a supervised injecting facility (SIF) located in Sydney, Australia. The aim was to gain a better understanding of the substances injected by clients through: (1) chemical analyses of the content of used syringes; (2) comparison of these results with clients' self-reported drug use; and (3) assessing the usefulness of analysing other injecting equipment to detect substances used. During one week in February 2019, syringes and other injecting equipment were collected at the Sydney SIF. Their residual content was analysed by gas-chromatography/mass-spectrometry. Heroin was the most commonly detected substance (present in 51% of syringes), followed by methamphetamine (22%) and oxycodone (10%). In addition to the main psychoactive substance, cutting agents reported in the literature were also detected in used syringes. The main psychoactive substance identified by laboratory analysis reliably corresponded with users' self-reported drug type. Analytical confirmation of substances injected allows for the provision of better targeted harm reduction messaging based on timely and objective data. The approach used is amenable to clients and feasible in the Australian SIF context. Upscaling and wider implementation could be done through Needle and Syringe Programs, and would support the early detection of harmful substances entering drug markets and better inform harm reduction strategies.

A rapid assay for the simultaneous determination of nicotine, cocaine and metabolites in meconium using disposable pipette extraction and gas chromatography-mass spectrometry (GC-MS).

Drug abuse by pregnant women is considered a serious public health problem worldwide. Meconium is the first excretion in newborns and has been used as an alternative matrix to evaluate in utero drug exposure. Solid phase extraction (SPE) is widely employed to prepare and clean up samples in the field of forensic analysis. Most SPE products require large volumes of solvent, which culminates in longer sample processing times and increased cost per sample. Disposable pipette extraction (DPX) tips have been used as an alternative to traditional SPE cartridges. They combine efficient and rapid extraction with reduced solvent consumption. The purpose of this study was to develop and validate a method to determine nicotine, cotinine, cocaine, benzoylecgonine, cocaethylene and methyl ester anhydroecgonine in meconium using DPX and gas chromatography-mass spectrometry (GC-MS). Validation results indicated that extraction efficiency ranged 50-98%, accuracy 92-106%, intra-assay precision 4-12% and inter-assay precision 6-12%. Linear calibration curves resulted in R(2) values >0.99, limits of detection ranged from 2.5 to 15 ng/g and the limit of quantitation from 10 to 20 ng/g. The DPX-GC-MS method was shown to selectively analyze trace concentrations of drugs in meconium samples. Finally, the developed and validated method was applied to 50 meconium samples.