Noor Azuwa Hamdan, Rahizan Issa, Mohd Fairulnizal Md Noh and Noraziah Mohamad Zin (2012). Electrochemical technique using Methylene Blue with Pencil Graphite Electrode for optimum detection of Mycobacterium tuberculosis DNA. Current Research in Tubercolusis 4(1): 1-12
The analysis of nucleic acids recognition using genosensor provides a rapid, sensitive and inexpensive detection for infectious and genetic diseases, bacteria food contaminations, forensic and environmental research. A simple application of electrochemical biosensor for the alternative detection of Mycobacterium tuberculosis (M. tuberculosis) using Pencil Graphite Electrode (PGE) and Methylene Blue (MB) as electroactive intercalators via non-covalent attachment was developed. Synthetic oligonucleotides of M. tuberculosis which consisted of M. Probe, M. Target, M. Non-complementary and M. Mutation were used for DNA hybridization detection on PGE. Differential Pulse Voltammetry (DPV) was performed using a PalmSens Electrochemical Portable Apparatus controlled by a Pocket PC. Various parameters affecting the response of the signals were explored and optimized including M. Probe concentration, immobilization time and hybridization time of immobilized M. Probe, concentration and accumulation time of MB, as well as the concentration of M. Target. The results obtained after measurement showed that the voltammetric signal of MB before hybridization was higher compared to after hybridization. This indicates that MB has high affinity towards guanine bases. Differences in the signals of MB for hybridization between M. Probes and Polymerase Chain Reaction (PCR) amplified products were observed. The Relative Standard Deviation (RSD) for TB PCR positive sample, TB PCR positive control, TB PCR negative sample, TB PCR negative control and TB PCR blank were 11.5, 2.9, 22.8, 13.8 and 20.3%, respectively which can be applied for the detection of M. tuberculosis.