A highly sensitive nanopore platform for measuring RNase A activity.

Ribonuclease A (RNase A) plays significant roles in several physiological and pathological conditions and can be used as a valuable diagnostic biomarker for human diseases such as myocardial infarction and cancer. Hence, it is of great importance to develop a rapid and cost-effective method for the highly sensitive detection of RNase A. The significance of RNase A assay is further enhanced by the growing attention from the biotechnology and pharmaceutical industries to develop RNA-based vaccines and drugs in large part as a result of the successful development of mRNA vaccines in the COVID-19 pandemic. Herein, we report a label-free method for the detection of RNase A by monitoring its proteolytic cleavage of an RNA substrate in a nanopore. The method is ultra-sensitive with the limit of detection reaching as low as 30 fg per milliliter. Furthermore, sensor selectivity and the effects of temperature, incubation time, metal ion, salt concentration on sensor sensitivity were also investigated.