MethylMeter® is a sensitive MSRE-free and bisulfite-free DNA methylation detection technology for the quantitative measurement of the methylation level of specific DNA targets, even in challenging clinical samples, including FFPE tissue and bodily fluids.
Detailed procedures for the use of MethlyMeter are availabe here, in the June 2016 Journal of Epigenomics.
MethylMeter uses Ribomed’s proprietary paramagnetic bead bound Me-CpG-binding domain (MBD) protein, MethylMagnet, to separate methylated DNA from unmethylated DNA. [Step 1]. MethylMagnet takes advantage of the high specificity of the MBD2 methyl-CpG binding domain and its bias for clustered CpGs, which favors the analysis of CpG islands. MethylMeter’s detection sensitivity is achieved with a novel signal generation process called Abscription® (Abortive Transcription). Abscription involves the reiterative synthesis of short ‘abortive’ RNA transcripts by a proprietary RNA polymerase (Abscriptase®) that remains bound at an Abortive Promoter Cassette (APC) generating multiple copies of a specific, short Abortive transcript (Abscript).
After separation with MethylMagnet Kits, APCs are attached to the DNA in the methylated and unmethylated fractions in a process call CAPS (Coupled Abscription PCR Signaling), [Step 2]. The APCs are attached to the amplicons generated from the DNA targets in the methylated and unmethylated fractions, and the amount of each abscript produced is a measure of the amount of the original DNA target in each fraction and can be used to determine the percentage methylation for that target.
In the fluorescent MethylMeter assays (Format II), each Abscript binds to a quenched molecular beacon and is then extended with CAPS polymerase in a beacon-dependent reaction to convert the single-stranded hairpin form of the beacon to the double stranded form. This separates the fluorophore from the quencher on the beacon and generates a fluorescent signal. The signal amplification provided by Abscription allows detection of as little as 6 copies of DNA in 36 cycles without preamplification or nested PCR, reducing the nonspecific amplification associated with higher cycle numbers.Current MethylMeter tests available as Research Use Only products are listed below.
|MethylMeter® and CAPS ™ Kits – Format II (Fluorescent Detection)|
|Description: These are target or biomarker panel specific kits already containing target-specific CAPS primers. MethylMeter is used to measure DNA methylation and includes MethylMagnet and CAPS. Each MethylMeter kit contains reagent for analysis of 12 samples and calibrators, and CAPS reactions on both the methylated and unmethylated fractions for each of the genes/CpG islands included in the assay. Mutation detection only requires CAPS.|
|Single target kits|
|Catalog #||Kit Name||Description|
|MM-MGMT-I||MethylMeter-MGMT-I||6-O-Methylguanine-DNA Methyltransferase promoter (downstream)|
|MM-MGMT-II||MethylMeter-MGMT-II||6-O-Methylguanine-DNA Methyltransferase promoter (upstream)|
|MM-DAPK1||MethylMeter-DAPK1||Death-Associated Protein Kinase 1|
|MM-hMLH1||MethylMeter-hMLH1||MutL Homolog 1 promoter|
|MM-RBI||MethylMeter-RBI||Retinoblastoma 1 imprinted|
|MM-SOWAHA||MethylMeter-SOWAHA||Sosondowah Ankyrin Repeat Domain Family Member A|
|MM-MAL||MethylMeter-MAL||T-Lymphocyte Maturation-Associated Protein|
|MM-SNRPN||MethylMeter-SNRPN||Small Nuclear Ribonucleoprotein Polypeptide N|
|CK-IDH1 R132H||CAPS-IDH1 WT and R132H detection||IDH1 R132H is primary mutation causing G-CIMP+|
|MM-GCIMP||MethylMeter- G-CIMP Kit||Determine G-CIMP (Glioma CpG Island Methylator Phenotype)|
|MM-GliomaSTRAT||MethylMeter- GliomaSTRAT Kit||Determine G-CIMP, MGMT methylation and IDH1 WT & R132H mutation|