Engineered NS1 for Sensitive, Specific Zika Virus Diagnosis from Patient Serology.

Dengue virus (DENV) and Zika virus (ZIKV) belong to the Flaviviridae family of viruses spread by Aedes aegypti mosquitoes in tropical and subtropical areas. Accurate diagnostic tests to differentiate the 2 infections are necessary for patient management and disease control. Using characterized ZIKV and DENV patient plasma in a blind manner, we validated an ELISA and a rapid immunochromatographic test for ZIKV detection. We engineered the ZIKV nonstructural protein 1 (NS1) for sensitive serologic detection with low cross reactivity against dengue and developed monoclonal antibodies specific for the ZIKV NS1 antigen. As expected, the serologic assays performed better with convalescent than acute plasma samples; the sensitivity ranged from 71% to 88%, depending on the performance of individual tests (IgM/IgG/NS1). Although serologic tests were generally less sensitive with acute samples, our ZIKV NS1 antibodies were able to complement the serologic tests to achieve greater sensitivity for detecting early infections.

CRISPR-based systems for sensitive and rapid on-site COVID-19 diagnostics.

The COVID-19 pandemic has strained healthcare systems. Sensitive, specific, and timely COVID-19 diagnosis is crucial for effective medical intervention and transmission control. RT-PCR is the most sensitive/specific, but requires costly equipment and trained personnel in centralized laboratories, which are inaccessible to resource-limited areas. Antigen rapid tests enable point-of-care (POC) detection but are significantly less sensitive/specific. CRISPR-Cas systems are compatible with isothermal amplification and dipstick readout, enabling sensitive/specific on-site testing. However, improvements in sensitivity and workflow complexity are needed to spur clinical adoption. We outline the mechanisms/strategies of major CRISPR-Cas systems, evaluate their on-site diagnostic capabilities, and discuss future research directions.

Levitational Cell Cytometry for Forensics.

Here, a method for label-free, real-time interrogation, monitoring, detection, and sorting of biological rare cells in magnetically suspended heterogeneous samples is developed. To achieve this, heterogeneous populations of cells are levitated and confined in a microcapillary channel. This strategy enables spatiotemporal differential magnetic levitation of rare fragile dead cells equilibrating at different heights based on the balance between magnetic and corrected gravitational forces. In addition, the sorting of fragile rare dead cell populations is monitored in real-time. This technique provides a broadly applicable label-free tool for high resolution, real-time research, as well as forensic evidence processing of rape kits. This method is validated with forensic mock samples dating back to 2003, isolating sperm from epithelial cells (E. cells) with >90% efficiency and >97% purity. Overall, this method reduces the processing time by over 20-fold down to 20 min, eliminating centrifugation and labels, and providing an inexpensive and high-yield alternative to the current centrifuge-based differential extraction techniques. It can potentially facilitate the forensic downstream genomic analyses, accelerating the identification of suspects, and advancing public safety.