Global DNA Methylation Assay–LINE-1 Kit

Global DNA Methylation Assay–LINE-1 Kit

Code: 55017

Global DNA Methylation – LINE-1 Kit Overview

Active Motif's Global DNA Methylation – LINE-1 Kit is an ELISA-based assay for the detection and quantification of global DNA methylation (i.e., 5-methylcytosine (5-mC) levels) in human genomic DNA. The assay measures 5-mC levels of Long Interspersed Nucleotide Element 1 (LINE-1) repeats. LINE-1 repeats represent approximately 17-18% of the human genome and have a copy number of approximately half a million. It is well established in the literature that the methylation status of these LINE-1 repeats serve as a surrogate measure of global DNA methylation.

Because changes in genomic DNA methylation levels are observed in various human diseases, measuring global DNA methylation has become an essential research method. Aberrant LINE-1 methylation is linked to the onset and poor prognosis of various diseases, including cancer. The 96-well Global DNA Methylation – LINE-1 Kit is ideally suited for screening variances in 5-mC levels across different disease states, treatment conditions, clinical outcomes, or environmental factors.

In the Global DNA Methylation – LINE-1 Assay, the genomic DNA of interest is first enzymatically digested using the provided MseI enzyme to generate the appropriate fragments to hybridize to a biotinylated consensus sequence corresponding to human LINE-1 transposon. The LINE-1 probe is designed to assay a 290 bp region of the LINE-1 repeat element containing 88 cytosine residues, of which 12 are in a CpG context. Hybridized samples are immobilized to a 96-well plate and methylated cytosines are identified using a 5-methylcytosine antibody, HRP-conjugated secondary antibody and colorimetric detection reagents. Generating a standard curve using the included DNA standards with known LINE-1 methylation levels provides the relative level of 5-methylcytosine in each DNA sample.

Global DNA Methylation – LINE-1 Kit Contents

The Global DNA Methylation – LINE-1 Kit contains sufficient reagents to perform 96 reactions. It is shipped on dry ice and contains reagents with multiple storage temperatures inside. Please confirm receipt of all reagents upon arrival and store items at the appropriate temperatures as indicated below. All reagents are guaranteed stable for 6 months from date of receipt when stored properly. This kit includes the following components:

  • LINE-1 Probe (2.5 μM); Store at -20°C

  • MseI enzyme (10 U/μl); Store at -20°C

  • 10X MseI Reaction Buffer; Store at -20°C

  • Methylated DNA Standard (10 ng/μl); Store at -20°C

  • Non-methylated DNA Standard (10 ng/μl); Store at -20°C

  • 5-Methylcytosine mAb; Store at -20°C

  • Anti-mouse HRP-conj. secondary Ab; Store at 4°C

  • Assay Buffer AM3; Store at 4°C

  • 20X Wash Buffer; Store at 4°C

  • 96-well Streptavidin plate; Store at 4°C

  • Developing Solution; Store at 4°C

  • Stop Solution; Store at RT

  • 5X Hybridization Buffer; Store at RT

  • 10X Buffer W; Store at RT

  • Plate sealer; Store at RT

  • 0.2 ml PCR stripwell tubes; Store at RT.

Global DNA Methylation – LINE-1 Kit Highlights

  • Compatibility: Use with cell or tissue derived human genomic DNA.

  • Sensitivity: Can detect a little as 0.5% methylation.

  • Input: Can detect 5-mC from as little as 10 ng of input DNA.

  • Quantitative: Included DNA standards contain known levels of LINE-1 methylation which can be used to determine % 5-mC.

  • Quick and Easy: Simple ELISA-based assay can be finished in 5 hours.