Design, synthesis, and application of electroactive reporters for nucleic acid biosensing
This thesis discusses the design, synthesis, and application of electroactive reporters for nucleic acid biosensing. The reporters are able to specifically recognize and sensitively report the hybridization of target nucleic acids to capture probes in an electrochemical biosensor. Each molecule contains a DNA intercalating moiety, 1,4,5,8naphthalenetetracarboxylic diimide, as the recognition unit that binds to nucleic acid duplexes. Four classes of compounds were studied: symmetrical mono-and
... metrical mono-and bisintercalators linked to either Os or Ru metal complex, symmetrical mono-intercalators linked to EDOT, and an asymmetrical mono-intercalator linked to both Os complex and EDOT. A range of redox potentials was obtained by modifying the ligands of the metal complexes, while binding properties was enhanced by positive charge. Three compounds were used in different detection schemes. In each scheme, direct detection was achieved by measuring the electroactivity of Os or Ru complex. Lower detection limits were obtained through various amplification strategies. The Os complex catalytically oxidizes ascorbic acid, leading to an amplified current signal in amperometry and a detection limit of 60 fM for oligonucleotide samples. The Ru complex catalytically oxidizes guanine bases, leading to an increase of peak current in cyclic voltammetry and a detection limit of 1.5 pM for TP53 cDNA. The EDOT moiety acts as 'seeds' that facilitates selective polymerization of poly(EDOT). Under controlled conditions, the amount of polymer deposited was positively related to target DNA concentrations as low as 20 pM. In conclusion, the advantageous electrochemical and binding properties of the newly synthesized reporters qualify them for application in electrochemical detection platforms that can potentially be incorporated into point-of-care nucleic acid biosensors.