DNA Biosensors for Clinical Diagnostics
Our early work in this area involved electrochemistry at DNA-modified electrodes. We found that single-stranded DNA could be covalently attached to oxidized carbon electrode surfaces, since these possess carboxylic acid groups that react with carbodiimides and N-hydroxysulfosuccinimide to form the NHS-ester. The deoxyguanosine residues of single-stranded DNA react with the NHS-esters to covalently bind the DNA to the electrode surface. Electrochemical methods, such as cyclic or differential pulse voltammetry, can be performed before and after this ss-DNA-modified electrode is allowed to react with the complementary DNA sequence. Solutions that contain redox-active transition metal polypyridine complexes, like Co(bpy)33+ , when examined with the DNA-modified electrode, show an enormous increase in voltammetric peak heights when the surface-bound DNA hybridizes with its complement to form surface-bound, double-stranded DNA. This method was applied to the detection of one of the mutations commonly found in cystic fibrosis patients and carriers. This mutation involves the deletion of three consecutive bases that code for a phenylalanine residue in the CF Transmembrane Regulatory Protein.
We are now working in collaboration with Clinical Microsensors, Inc., of Pasadena, CA, to develop electrochemical DNA biosensors based on miniature gold electrodes. The DNA used to modify the electrode has been synthesized to contain a terminal thiol group, which chemisorbs to the gold electrode to form a reasonably stable linkage. These electrodes are being studied electrochemically and using our new Atomic Force Microscope, which can provide atomic-resolution underwater images of their surfaces. Our target diagnostic applications now involve disorders caused by abnormally high repeat numbers of three-base sequences, such as Huntington's disease and Fragile X syndrome.
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