Table 4.
Methods of detection of DNA methylation in circulating cells
| Detection type | Method | Description | Refs. |
|---|---|---|---|
| Site-specific detection | Conventional MSP | Requiring a sample spot (5 ml of peripheral blood); Able to be used in the detection of certain methylated genes in the plasma of serum; using specific PCR primers for methylated sequences | [156], [157], [158] |
| Fluorescence-based real-time MSP | Facilitating quantitative detection; sensitive; requiring prior knowledge of the methylated sequences | [174] | |
| QDs-FRET | Able to reduce the background for detecting targets at low concentration; greater sensitivity; limited FRET efficiency; impractical for challenging samples such as serum and plasma | [163] | |
| MOB | Easy to handle; increased detection throughput; providing efficient, sensitive methylation detection in diagnosis; able to be used in blood samples | [160], [164] | |
| cMethDNA | High sensitivity, specificity, reproducibility, dynamic range, and quantitative advantages; detecting methylated site at low levels in cell-free circulating serum DNA; promising new liquid biopsy tool | [165] | |
| Genome-scale detection | Conventional bisulfite conversion-based methods | Gold standard for the detection of DNA methylation; requiring a relatively large amount of sample; focused on CpG islands or promoter regions | [166], [167], [168] |
| Conventional enrichment-based methods | No conversion treatment; requiring a high concentration of DNA; likely ignoring other methylated sites when using antibody against 5 mC or 5 mCG | [169], [170], [171] | |
| Short-gun massively parallel bisulfite sequencing | Detecting with high sensitivity and specificity even at a low sequence depth with 10 million sequencing data; requiring 4 ml plasma only | [172] | |
| MCTA-seq | Working well with ctDNA samples as small as 7.5 pg; able to simultaneously detect thousands of hypermethylated CpG islands in cfDNA | [173] |
Note: MSP, methylation-specific PCR; QDs-FRET, quantum dots-fluorescence resonance energy transfer; MOB, methylation on beads; MCTA-seq, methylated CpG tandem amplification and sequencing; ctDNA, circulating tumor DNA; cfDNA, cell-free DNA.