DNA Methylation Readers in Biosensing Strategies

DNA methylation is an epigenetic modification that influences gene expression and cellular signaling. Abnormalities in the DNA methylation landscape have been associated with diverse pathologies and diseases, making accurate and accessible detection methods essential for both research and clinical diagnostics. Conventional methylation analysis techniques, such as bisulfite conversion-based assays, are the gold standard; however, they can be labor-intensive, chemically harsh, and poorly suited for real-time or point-of-care applications. Additionally, the potential for incomplete or over-conversion may affect diagnostic performance and has been a longstanding topic of discussion around bisulfite conversion. Alternative strategies, such as methylation-sensitive restriction enzymes (MSREs) and catalytic approaches, have addressed some limitations but still face challenges in specificity and sensitivity, throughput, and operational complexity. Recent advances in bioengineering and nanotechnology have leveraged the development of molecular elements, broadly referred to as "methylation readers", capable of specifically recognizing methylated DNA. Methylation readers are naturally occurring proteins, such as methyl-CpG binding domain proteins (MBDs), but also synthetically engineered proteins, such as transcription activator-like effectors (TALEs), as well as abiotic analogs, such as molecularly imprinted polymers (MIPs) and methylation-sensitive intercalators. The recent integration of these methylation readers into various biosensor platforms has enabled a significant advancement in epigenetic analysis, offering rapid, sensitive, and conversion-free DNA methylation detection without the need for relatively complex sample preparation. Such capabilities hold significant potential for early disease detection, personalized diagnostics, and fast monitoring of methylation-associated diseases. This review aims to provide a comprehensive and focused overview of the recent developments in the integration of methylation readers into biosensing technologies. We highlight the potential of these integrated approaches to transform epigenetic analysis in both research and clinical settings, discuss existing limitations, and outline future directions for advancing the field.