Reza Nouri

The outbreak of the SARS-CoV-2 caused the disease COVID-19 to spread globally. Specific and sensitive detection of SARS-CoV-2 facilitates early intervention and prevents the disease from spreading. Here, we present a solid-state CRISPR-Cas12a-assisted nanopore (SCAN) sensing strategy for the specific detection of SARS-CoV-2. We introduced a nanopore-sized counting method to measure the cleavage ratio of reporters, which is used as a criterion for positive/negative classification. A kinetic cleavage model was developed and validated to predict the reporter size distributions. The model revealed the trade-offs between sensitivity, turnaround time, and false-positive rate of the SARS-CoV-2 SCAN. With preamplification and a 30 min CRISPR Cas12a assay, we achieved excellent specificity against other common human coronaviruses and a limit of detection of 13.5 copies/μL (22.5 aM) of viral RNA at a confidence level of 95%. These results suggested that the SCAN could provide a rapid, sensitive, and specific analysis of SARS-CoV-2.

Quantification of HIV RNA in plasma is critical for identifying the disease progression and monitoring the effectiveness of antiretroviral therapy. While RT-qPCR has been the gold standard for HIV viral load quantification, digital assays could provide an alternative calibration-free absolute quantification method. Here, we reported a Self-digitization Through Automated Membrane-based Partitioning (STAMP) method to digitalize the CRISPR-Cas13 assay (dCRISPR) for amplification-free and absolute quantification of HIV-1 viral RNAs. The HIV-1 Cas13 assay was designed, validated, and optimized. We evaluated the analytical performances with synthetic RNAs. With a membrane that partitions ∼100 nL of reaction mixture (effectively containing 10 nL of input RNA sample), we showed that RNA samples spanning 4 orders of dynamic range between 1 fM (∼6 RNAs) to 10 pM (∼60k RNAs) could be quantified as fast as 30 min. We also examined the end-to-end performance from RNA extraction to STAMP-dCRISPR quantification using 140 μL of both spiked and clinical plasma samples. We demonstrated that the device has a detection limit of approximately 2000 copies/mL and can resolve a viral load change of 3571 copies/mL (equivalent to 3 RNAs in a single membrane) with 90% confidence. Finally, we evaluated the device using 140 μL of 20 patient plasma samples (10 positives and 10 negatives) and benchmarked the performance with RT-PCR. The STAMP-dCRISPR results agree very well with RT-PCR for all negative and high positive samples with Ct < 32. However, the STAMP-dCRISPR is limited in detecting low positive samples with Ct > 32 due to the subsampling errors. Our results demonstrated a digital Cas13 platform that could offer an accessible amplification-free quantification of viral RNAs. By further addressing the subsampling issue with approaches such as preconcentration, this platform could be further exploited for quantitatively determining viral load for an array of infectious diseases.

BACKGROUND: The aim of this study was to investigate the effects of exercise training on maximum aerobic capacity, resting heart rate (RHR), blood pressure and anthropometric variables of postmenopausal women with breast cancer. METHODS: Twenty nine women with breast cancer who received surgery, chemotherapy and radiotherapy with current hormone therapy were divided into two groups; intervention and control. Subjects in the intervention group performed 15 weeks combination exercise training including walking for 25 to 45 minutes (2 sessions per week) and resistance training for 60 minutes (2 sessions per week that were different from walking days). In pre and post tests, VO(2)max, RHR, blood pressure, body weight, body mass index (BMI) and waist to hip ratio (WHR) were measured in both groups. Data was analyzed using analysis of covariance (ANCOVA). RESULTS: Significant differences were observed for VO (2)max, RHR, body weight, BMI and WHR between intervention and control groups after 15 weeks (p < 0.05). In fact, exercise training had positive effects on the VO (2)max, RHR, body weight, BMI and WHR in postmenopausal women with breast cancer. No significant different was found for blood pressure between two groups (p > 0.05). CONCLUSIONS: It can be concluded that exercise training may improve maximum aerobic capacity, RHR and anthropometric variables in postmenopausal women with breast cancer.