Studies on the Purification of Recombinant Protein Biomarkers and the Development of Aptamers against Cardiac Troponin T (cTnT)

Abstract

For several decades, researchers have attempted to find aptamers for biomarkers because of their importance for the diagnosis of disease, biosensing and drug targeting. An aptamer is a powerful biological ligand due to the integrity of its extraordinary binding affinity and discrimination to specific target molecules. Cardiac troponin T (cTnT), which is the tropomyosin-binding subunit, is a reliable biomarker for the early diagnosis of acute myocardial infarction (AMI). Neutrophil gelatinase-associated lipocalin (NGAL), which is involved in innate immunity by sequestrating iron that, in turn, limits bacterial growth, is trustworthy biomarker for acute kidney injury (AKI). In this study, both recombinant proteins cTnT and NGAL as certain biomarkers are successfully developed. Research on cTnT structure has been insufficient owing to its poor solubility and obvious tendency to aggregate. Although cTnT is an unstable protein, the use of the dialysis tool made purification possible. And then, single-stranded deoxyribonucleic acid (ssDNA) aptamer candidates against cTnT were developed using the systematic evolution of ligands by exponential enrichment (SELEX) method. Among several SELEX techniques, such as affinity chromatography, capillary electrophoresis, and the magnetic beads-based method, the magnetic beads-based strategy was used because of its simplicity and quickness. The developed aptamers are expected to have many potential applications as a bioprobe for AMI.