Combination-Lock SlipChip Integrating Nucleic Acid Sample Preparation and Isothermal LAMP Amplification for the Detection of SARS-CoV-2

Abstract

Nucleic acid analysis with an easy-to-use workflow, high specificity and sensitivity, independence of sophisticated instruments, and accessibility outside of the laboratory is highly desirable for the detection and monitoring of infectious diseases. Integration of laboratory-quality sample preparation on a hand-held system is critical for performance. A SlipChip device inspired by the combination lock can perform magnetic bead-based nucleic acid extraction with several clockwise and counterclockwise rotations. A palm-sized base station was developed to assist sample preparation and provide thermal control of isothermal nucleic acid amplification without plug-in power. The loop-mediated isothermal amplification reaction can be performed with a colorimetric method and directly analyzed by the naked eye or with a mobile phone app. This system achieves good bead recovery during the sample preparation workflow and has minimal residue carryover from the lysis and elution buffers. Its performance is comparable to that of the standard laboratory protocol with real-time qPCR amplification methods. The entire workflow is completed in less than 35 min and the device can achieve 500 copies/mL sensitivity. Thirty clinical nasal swab samples were collected and tested with a sensitivity of 95% and a specificity of 100% for SARS-CoV-2. This combination-lock SlipChip provides a promising fast, easy-to-use nucleic acid test with bead-based sample preparation that produces laboratory-quality results for point-of-care settings, especially in home use applications.

CRISPR/Cas12a-Powered Amplification-Free RNA Diagnostics by Integrating T7 Exonuclease-Assisted Target Recycling and Split G-Quadruplex Catalytic Signal Output

“…RNAs, as genetic massagers in biological cells, play an irreplaceable role in gene coding and regulation across almost all life processes. , Its great value in disease diagnosis and prognosis assessment has also attracted considerable interest in recent years . For instance, bacterial 16S rRNA (rRNA), which is highly conserved between different bacterial species and relatively abundant in viable cells, serves as an ideal target in species-specific pathogen identification. − Noncoding RNAs, including MicroRNAs and long noncoding RNA (lncRNA), are another important kind of RNA that participate in life activities as “organizers” and regulatory molecules, and their aberrant expression has become an important diagnostic and prognostic biomarker for malignant tumors, cardiovascular diseases, and nervous system diseases. , Although traditional methods, such as reverse transcription-quantitative polymerase chain reaction (RT-qPCR), Northern blots, and microarray analysis, have been well established for RNA detection, the involvement of tedious process, dedicated equipment, and trained personnel largely limited their applications to centralized laboratories. , Isothermal amplification techniques, including nucleic acid sequence-based amplification (NESBA), recombinase polymerase amplification (RPA), and loop-mediated isothermal amplification (LAMP), avoid complex thermal cycling and shorten the detection time. − However, their applications are still restricted by the sophisticated primer design and the risk of aerosol contamination. Therefore, a rapid, simple, and sensitive method for RNA detection remains an urgent need.…”