In Vitro Selection of a DNA Aptamer Targeting Degraded Protein Fragments for Biosensing
Meng Liu, Jiayi Wang, Yangyang Chang, Qiang Zhang, Dingran Chang, Christy Y. Hui, John D. Brennan, Yingfu Li
Abstract
Protein biomarkers often exist as degradation fragments in biological samples, and affinity agents derived using a purified protein may not recognize them, limiting their value for clinical diagnosis. Herein, we present a method to overcome this issue, by selecting aptamers against a degraded form of the toxin B protein, which is a marker for diagnosing toxigenic Clostridium difficile infections. This approach has led to isolation of a DNA aptamer that recognizes degraded toxin B, fresh toxin B, and toxin B spiked into human stool samples. DNA aptamers selected using intact recombinant toxin B failed to recognize degraded toxin B, which is the form present in stored stool samples. Using this new aptamer, we produced a simple paper-based analytical device for colorimetric detection of toxin B in stool samples, or in the NAP1 strain of Clostridium difficile. The combined aptamer-selection and paper-sensing strategy can expand the practical utility of DNA aptamers in clinical diagnosis.