Aptamer-based cell-free detection system to detect target protein.

Biomarkers of disease, especially protein, show great potential for diagnosis and prognosis. For detecting a certain protein, a binding assay implementing antibodies is commonly performed. However, antibodies are not thermally stable and may cause false-positive when the sample composition is complicated. In recent years, a functional nucleic acid named aptamer has been used in many biochemical analysis cases, which is commonly selected from random sequence libraries by using the systematic evolution of ligands by exponential enrichment (SELEX) techniques. Compared to antibodies, the aptamer is more thermal stable, easier to be modified, conjugated, and amplified. Herein, an Aptamer-Based Cell-free Detection (ABCD) system was proposed to detect target protein, using epithelial cell adhesion molecule (EpCAM) as an example. We combined the robustness of aptamer in binding specificity with the signal amplification ability of CRISPR-Cas12a's -cleavage activity in the ABCD system. We also demonstrated that the ABCD system could work well to detect target protein in a relatively low limit of detection (50-100 nM), which lay a foundation for the development of portable detection devices. This work highlights the superiority of the ABCD system in detecting target protein with low abundance and offers new enlightenment for future design and development.