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Fast and accurate detection of Cryptococcus and precise differentiation of its subtypes is of great significance in protecting people from cryptococcal disease and preventing its spread in populations. However, traditional Cryptococcus identification and detection techniques still face significant challenges in achieving high analysis speed as well as high sensitivity. In this work, we report an electric microfluidic biochip. Compared to conventional methods that take several hours or even a day, this chip can detect Cryptococcus within 20 min, and achieve its maximum detection limit within 1 h, with the ability to differentiate between the Cryptococcus neoformans (NEO) and rare Cryptococcus gattii (GAT) efficiently, which accounts for nearly 100%. This device integrated two functional zones of an electroporation lysis (EL) zone for rapid cell lysis (<30 s) and an electrochemical detection (ED) zone for sensitive analysis of the released nucleic acids. The EL zone adopted a design of microelectrode arrays, which obtains a large electric field intensity at the constriction of the microchannel, addressing the safety concerns associated with high-voltage lysis. The device enables a limit of detection (LOD) of 60 pg/mL for NEO and 100 pg/mL for GAT through the modification of nanocomposites and specific probes. In terms of the detection time and sensitivity, the integrated microfluidic biochip demonstrates broad potential in Cryptococcus diagnosis and disease prevention. [ABSTRACT FROM AUTHOR]

Precise quantification of low-dose ionizing radiation is of great significance in protecting people from damage caused by clinical radiotherapy or environmental radiation. Traditional techniques for detecting radiation, however, remain extreme challenges to achieve high sensitivity and speed in quantifying radiation dosage. In this work, we report a Cas13a-Microdroplet platform that enables sensitive detection of ultra-low doses of radiation (0.5 Gy vs. 1 Gy traditional) within 1 h. The micro-platform adopts an ideal, specific radiation-sensitive marker, m6A on NCOA4 gene (NCOA4-m6A) that was first reported in our recent work. Microfluidics of the platform generate uniform microdroplets that encapsulate a CRISPR/Cas13a detection system and NCOA4-m6A target from the whole RNA extraction, achieving 10-fold enhancement in sensitivity and significantly reduced limit of detection (LOD). Systematic mouse models and clinical patient samples demonstrated its superior sensitivity and LOD (0.5 Gy) than traditional qPCR, which show wide potentials in radiation tracking and damage protection. [ABSTRACT FROM AUTHOR]

Probing nuclear protein expression while correlating cellular behavior is crucial for deciphering underlying causes of cellular disorders, such as tumor metastasis. Despite efforts to access nuclear proteins by trafficking the double barriers of cell membrane and nuclear membrane, they mostly fall short of the capacity for analyzing various proteins in different cells. Herein, we introduce a Companion-Probe & Race (CPR) platform that enables interrogating nuclear proteins in living cells, while guiding and tracking cellular behaviors (e.g., migration) in real time. The Companion-Probe consists of two polypeptide complexes that were structured with nuclear localization signal (NLS) for entering nucleus, recognition polypeptide for targeting different sites of nuclear proteins, and fragments of green fluorescent protein (GFP) that can recover a whole fluorescent GFP once the two polypeptide complexes combine with a same target protein. The two polypeptide complexes were expressed by two plasmids (named "probe plasmids") that were uniformly and efficiently delivered into cells by nano-electroporation (NEP), a high-performance delivery method for cell focal-poration and accelerated intracellular delivery. To track cell migration, multiple radial microchannels were designed with micro-landmarks on the platform to serve as addressable runways for cells. The proof-of-concept of CPR platform was validated with clinical primary cells that indicated the positive-correlation between nuclear protein murine double minute 2 (MDM2) expression level and cell migration velocity. This platform shows great promises to interrogate nuclear proteins in live cells, and to decode their roles in determining cellular behaviors on a chip. [ABSTRACT FROM AUTHOR]