TABLE 1.

Comparison of selected methods for DNAm analysis.

Method	Mechanism for DNAm detection	Genome wide analysis	Targeted DNAm analysis	Accuracy of DNAm at individual CpGs	Integrative analysis with other datasets	Scalability for many samples	CE accredited instrumentation a	Bioinformatic requirement	Usual turn arround time	Costs b
Whole Genome Bisulfite Sequencing (WGBS)	High-throughput sequencing of bisulfite or enzym. converted DNA	++	—	+	++	-	+	↑↑	↑↑	↑↑
Reduced Representation Bisulfite Sequencing (RRBS)	High-throughput sequencing of bisulfite or enzym. converted DNA	+	—	+	+	+	—	↑↑	↑↑	↑
Infinium BeadChip Technology (EPIC array)	Microarray analysis of bisulfite-converted DNA	+	—	++	++	+	—	↑	↑	↑
Nanopore sequencing	Detection of current changes during sequencing of non-converted DNA	++	—	-	+	+	+	↑↑	↑	↑
Methylated DNA immunoprecipitation (MeDIP)	Affinity capture of unconverted DNA with antibody and deep sequencing	++	NA	-	+	-	—	↑↑	↑↑	↑
Methyl Binding Domain (MBD) Sequencing	Affinity capture of unconverted DNA with MBD and deep sequencing	++	NA	-	+	-	—	↑↑	↑↑	↑
Probabilistic frameworks for genome wide DNAm pattern analysis	Shallow sequencing data (e.g. RRBS for scAge or methyl-ATAC-seq for TIME-seq)	-		-	+	++	—	↑↑	↑↑	↔
Methylation sensitive restriction enzyme (MSRE) PCR	Unconverted DNA is digested with MRSE and amplified by PCR	NA	+	-	-	+	—	-	↔	↓
Methylation specific PCR	PCR amplicons of bisulfite-converted or enzymatic converted DNA	NA	+	-	-	+	—	-	↔	↓
Quantitative PCR (qPCR)	Allele specific qPCR after bisulfite-conversion	NA	+	+	-	+	—	-	↔	↓
Targeted amplicon sequencing	Deep-sequencing of PCR amplicons of bisulfite converted DNA	NA	+	++	-	++	+	↑	↑	↔
Pyrosequencing	Sequencing of PCR amplicons of bisulfite-converted DNA	NA	+	++	-	+	—	-	↔	↔
Digital droplet PCR (ddPCR)	PCR of bisulfite-converted DNA in droplets	NA	+	++	-	+	+	-	↔	↔
EpiTYPER	Mass-spectrometry of bisulfite-converted DNA	NA	+	++	-	+	—	-	↔	↔

This table shall only exemplify how hallmarks of DNAm biomarkers may be influenced by different methods. It does not claim to represent all available approaches for DNAm analysis. The suggested classifications will vary between applications and laboratories.

^aAccreditation may also include enzymes and viable components. The regulatory requirements and accreditation may change with time and according to local regulations.

^bThe expenses can only be estimated relatively. They include consumables, instrumentation and personnel costs and depend largely on the number of samples that can be processed in parallel (and on the number of CpGs for the targeted assays). NA, not applicable.