Epiontis applies different assay methods for quantitative and highly sensitive analysis of therapeutic cells.
For quantitative cell characterization, Epiontis uses a proprietary methylation-specific single base extension procedure (MS-SNuPE). Individual, predictive cytosines in differentially methylated regions are analysed. Upon chemical conversion the DNA is used as template for specific primer extension,with the primer designed to anneal directly adjancent to the Cytosine of interest. Chain elongation is terminated directly after adding the first base, since fluorescently labeled ddNTPs serve as substrate. Due to the primer design, this base is always correspondent to either a methylated or unmethylated cytosine. Mixtures of signal represent the ratio of cells with methylated versus unmethylated Cytosine at the analysed position.
When the detetion of minute amounts of a cell fraction/potential contamination is required, Epiontis uses its proprietary, highly sensitive detection methods. Our methods potentially allow the detection of a single copy of DNA. They are based on real time PCR and specifically amplify either methylated or unmethylated DNA. For example, in an overwhelmingly unmethylated pool of DNA, individual methylated copies can be specifically detected and vice versa. A PCR product is generated using methylation unspecific primers, at the same time a blocker specifically inhibits the amplification of one of the possible alleles from amplification. With this high sensitivity and excellent specificity is achieved.
