Development of a Molecular Diagnosis Assay Based on Electrohybridization at Plastic Electrodes and Subsequent PCR
Reliable recognition of specific DNA sequences will play a leading role in future medical diagnostic procedures. Whereas many approaches to molecular diagnosis systems include DNA microarrays on chips and fluorometric detection, the basis of our approach is the use of inexpensive components, especially plastic electrodes, with low multiplexing and an electrochemical detection unit. To increase the sensitivity, PCR can be used as an intermediate step.
For selective enrichment, the 5?-end of specific nucleic acid probes have been attached covalently to free carboxyl groups of conducting polycarbon/ carbon fiber electrodes.
Specific oligonucleotides have been enriched by cyclic inversion of an electrochemical potential, transferred into a PCR vial and thermally desorbed, optionally supported by electrochemical repulsion. The analysis of the PCR product shows the efficiency and selectivity of the electrochemical enrichment.
Hybridization of DNA can be shown by different electrochemical methods often combined with electroactive markers like osmium tetroxide and daunomycin.
Recently we have shown that inexpensive and robust electrodes made of polycarbon and conductive carbon powder are suitable for electrochemical examination of nucleic acids. The oxidative transfer stripping guanine peak of DNA can easily be resolved at concentrations less than 1mg/ml DNA.
The combination of electrochemical enrichment of DNA and subsequent detection is a promising approach towards an inexpensive molecular diagnosis kit, optionally equipped with an additional PCR unit.
J. Schülein, B.Grassl, J.Krause, J.Hassmann, P.Müller1, W.M.Bertling
november AG, Ulrich-Schalk-Str. 3, 91056 Erlangen, Germany