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However, the use of specialized PCR equipment makes these approaches labor-intensive, time-consuming and quite expensive, thus limiting their use in general laboratories. However, they are typically prone to high variability and, therefore, only suitable for the rough estimation of DNA methylation Fluorescence, capillary electrophoresis 19 , colorimetry 20 , surface plasmon resonance 21 and surface enhanced Raman spectroscopy 22 have been also employed to detect specific DNA methylation pattern These methodologies, although effective, tend to be expensive, tedious and DNA and time-consuming for many applications.
In addition, the affinity of the methyl-binding protein MBD 24 , 25 and the anti-methylcytosine antibody antimC 4 , 14 for 5-mC in a double stranded ds – or single stranded ss -target DNA sequence, respectively, has been exploited to construct a few biosensors for rapid monitoring of DNA methylation 9 , However, these biosensors lack nucleic acid amplification, this constraining their sensitivity and applicability for the analysis of real specimens containing low amounts of methylated DNA.
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The applicability of these strategies has been demonstrated by targeting the promoter methylation status of two normally non-methylated, biologically significant cancer genes, both in spiked biological fluids RASSF1A , and genomic DNA extracted from human glioblastoma cells MGMT.
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