DNA methylation has been extensively investigated in recent years, not least because of its known relationship with various diseases. Progress in analytical methods can greatly increase the relevance of DNA methylation studies to both clinical medicine and scientific research. Microfluidic chips are excellent carriers for molecular analysis, and their use can provide improvements from multiple aspects. On-chip molecular analysis has received extensive attention owing to its advantages of portability, high throughput, low cost, and high efficiency. In recent years, the use of novel microfluidic chips for DNA methylation analysis has been widely reported and has shown obvious superiority to conventional methods. In this review, we first focus on DNA methylation and its applications. Then, we discuss advanced microfluidic-based methods for DNA methylation analysis and describe the great progress that has been made in recent years. Finally, we summarize the advantages that microfluidic technology brings to DNA methylation analysis and describe several challenges and perspectives for on-chip DNA methylation analysis. This review should help researchers improve their understanding and make progress in developing microfluidic-based methods for DNA methylation analysis.