Holiday Winter Sale Holiday Winter Sale: Use code E1712WNT to save 15% on your entire order. Ends 12/29/17. » Learn More
Epigentek Home
 
EpiGentek Guarantee

MethylFlash Methylated DNA Quantification Kit (Fluorometric)

Share:

For fluorescence-based quantitation of global DNA methylation in an ELISA-like format

Citations (26) | (1) | Write a Review
Suggested Workflow
DNA Isolation
 
 
DNA Methylation Quantification
 
Schematic procedure for the MethylFlash™ Methylated DNA Quantification Kit (Fluorometric).
Demonstration of high sensitivity and specificity of methylated DNA detection achieved by the MethylFlash™ kit. Synthetic unmethylated DNA (contains 50% of cytosine) and methylated DNA (contains 50% of 5-methylcytosine) were added into the assay wells at different concentrations and then measured with the MethylFlash™ Methylated DNA Quantification Kit (Fluorometric).
Input Type: DNA
Research Area: DNA Methylation
Target Application: Amount Quantitation
Vessel Format: 96-Well Plate
100% Guarantee: 6 months
Catalog No.SizePriceQty
P-1035-4848 assays $317.00 
P-1035-9696 assays $559.00 
Availability: Usually Ships in 1 Day or Same day delivery Same Day NY Delivery 
Product Overview

The MethylFlash™ Methylated DNA Quantification Kit (Fluorometric) is a complete set of optimized buffers and reagents to fluorometrically quantify global DNA methylation by specifically measuring levels of 5-methylcytosine (5-mC) in a microplate-based format. Compared to LUMA or LINE-1, Alu, and LTR-based assays, MethylFlash™ technology directly quantifies actual global DNA methylation.

This kit is also specifically optimized for paired use with the MethylFlash Hydroxymethylated DNA Quantification Kit (Fluorometric) for simultaneously quantifying both methylated DNA and hydroxymethylated DNA.

Product Features
The MethylFlash™ Methylated DNA Quantification Kit (Fluorometric) is a further refinement of our previous SuperSense™ global DNA methylation quantification kits by simplifying the workflow and improving the consistency of results. It uses an innovative method to enable background signals to be extremely low, eliminating the plate drying and blocking steps. Compared to chromatography-based methods such as HPLC or mass spectrometry, this product provides a cost-effective way to accurately measure levels of 5-methylcytosine. The kit has the following advantages and features:

  • Fluorometric assay with easy-to-follow steps for convenience and speed. The entire procedure can be finished within 4 hours.
  • Innovative kit composition enables background signals to be extremely low, which eliminates the need for plate blocking and allows the assay to be simple, accurate, reliable, and consistent.
  • High sensitivity, of which the detection limit can be as low as 50 pg of methylated DNA and accepts as low as 20 ng of input genomic DNA
  • Optimized antibody and enhancer solutions allow high specificity to 5-mC, with no cross-reactivity to unmethylated cytosine and no or negligible cross-reactivity to hydroxymethylcytosine within the indicated concentration range of the sample DNA.
  • Universal positive and negative controls are included, which are suitable for quantifying methylated DNA from any species.
  • Strip-well microplate format makes the assay flexible: manual or high throughput analysis.

Principle & Procedure
The MethylFlash™ Methylated DNA Quantification Kit (Fluorometric) contains all reagents necessary for the quantification of global DNA methylation. In this assay, DNA is bound to strip wells that are specifically treated to have a high DNA affinity. The methylated fraction of DNA is detected using capture and detection antibodies and then quantified fluorometrically by reading the absorbance in a fluorescence microplate spectrophotometer at excitation 530 and emission 590 nm. The amount of methylated DNA is proportional to the RFU (relative fluorescence units) measured, which can be calculated with the kit's included formulas for relative methylation status of two different DNA samples or absolute quantification of 5-methylcytosine (5-mC) using a standard curve.

Easy, Fast, and Flexible
The entire fluorometric assay has easy-to-follow steps for convenience and speed, which can be completed in 4 hours. The strip-well microplate format allows for a flexible assay in manual or high throughput analysis. Universal positive and negative controls are included with the kit for quantifying methylated DNA from any species such as mammals, plants, fungi, bacteria, and viruses.

Safe and Convenient
All the needed reagents, including negative controls and positive controls, for quantification of global DNA methylation are conveniently packaged in the kit. The direct fluorometric quantification of DNA samples replaces obsolete or inferior methods and eliminates the need for DNA digestion/denaturation, radioactivity, extraction, or chromatography.

Highly Sensitive and Specific
The novel procedure and proprietary kit compositions allow for accurate quantification of methylated DNA to be achieved with high sensitivity and specificity. The detection limit of the input DNA can be as low as 50 pg of methylated DNA.

Comparison of Available ELISA-based Global DNA Methylation Assay Methods 

 MethylFlash (Fluorometric)Competitor ZCompetitor C
Assay Principle Direct ELISA Direct ELISA Indirect ELISA
Format 96-well plate 96-well plate 96-well plate
Sensitivity Excellent detection limit: 0.05 ng of 5-mC DNA Good detection limit: >0.5 ng of 5-mC DNA Very poor detection limit: >1000 ng of 5-mC DNA
S/N Ratio >20 with very low background  <4 with high background  N/A*
Procedural Convenience No need for plate blocking and denaturation of DNA Requires plate blocking and DNA denaturation Requires plate blocking, DNA digestion, and antibody competitive incubation in an additional plate
Protocol Time  <4 hr  <4 hr  >8 hr
DNA Type & Species Universal for any species, both ssDNA and dsDNA Human and mouse ssDNA only Various species
Specificity Specific to 5-mC only Cross-reaction to 5-hmC N/A*
Quantitation Type Both relative and absolute quantification Relative Relative
Standard Control Stable, quantitative in absolute amount of 5-mC, and can be universally used for any species Unstable, cannot be quantitative in amount, and can only be used for human or mouse N/A*
Minimum Input Amount 20 ng 100 ng 1000 ng
Accuracy of Detection High, stable and quantified standard control. Proven close correlation with LC-MS analysis by users Unclear, unstable and un-quantified standard control N/A*
% 5-mC calculation Simple Complicated Not Provided
Patented Method Yes No No
Popularity Very high published citation count Low published citation count Low published citation count 
Support Expertise Since 2006 Since 2013 Since 2011

* Insufficient information provided by datasheet or product failed to produce data to make conclusive determination


Fig. 1. Schematic procedure for the MethylFlash™ Methylated DNA Quantification Kit (Fluorometric).


Fig. 2. Demonstration of high sensitivity and specificity of methylated DNA detection achieved by the MethylFlash™ kit. Synthetic unmethylated DNA (contains 50% of cytosine) and methylated DNA (contains 50% of 5-methylcytosine) were added into the assay wells at different concentrations and then measured with the MethylFlash™ Methylated DNA Quantification Kit (Fluorometric).

Product Components

MF1 (10X Wash Buffer)
MF2 (Binding Solution)
MF3 (Negative Control, 20 µg/ml)*
MF4 (Positive Control, 20 µg/ml)*
MF5 (Capture Antibody, 1000 µg/ml*
MF6 (Detection Antibody, 400 µg/ml)*
MF7 (Enhancer Solution)*
MF8 (Fluoro Developer)*
MF9 (Fluoro Enhancer)*
MF10 (Fluoro Dilutor)
8-Well Assay Strips (With Frame)
User Guide

* Spin the solution down to the bottom prior to use.

Note: The MF3 Negative Control is an unmethylated polynucleotide containing 50% of cytosine. The MF4 Positive Control is a methylated polynucleotide containing 50% of 5-methylcytosine.

User Guide & MSDS

[User Guide]*
*Always use the actual User Guide that shipped with your product. Is the above file locked? You can also request user guides by emailing info@epigentek.com along with your contact information and institution name.

Product Citations

Arena R et. al. (November 2017). Developmental peculiarities in placentae of ovine uniparental conceptuses. PLoS One. 12(11):e0188278.

Zhu Y et. al. (October 2017). Maternal dietary manganese protects chick embryos against maternal heat stress via epigenetic-activated antioxidant and anti-apoptotic abilities. Oncotarget. 8(52):89665-89680.

Geyer KK et. al. (May 2017). The Biomphalaria glabrata DNA methylation machinery displays spatial tissue expression, is differentially active in distinct snail populations and is modulated by interactions with Schistosoma mansoni. PLoS Negl Trop Dis. 11(5):e0005246.

Yue Ma et. al. (April 2017). Titanium dioxide nanoparticles induce size-dependent cytotoxicity and genomic DNA hypomethylation in human respiratory cells Royal Society of Chemistry. 2017(7):23560-23572.

Seo JS et. al. (February 2017). Hinokitiol induces DNA demethylation via DNMT1 and UHRF1 inhibition in colon cancer cells. BMC Cell Biol. 18(1):14.

Wang R et. al. (February 2017). Basic studies on epigenetic carcinogenesis of low-dose exposure to 1-trichloromethyl-1,2,3,4-tetrahydro-β-carboline (TaClo) in vitro. PLoS One. 12(2):e0172243.

Adamu HA et. al. (January 2017). In utero exposure to germinated brown rice and its oryzanol-rich extract attenuated high fat diet-induced insulin resistance in F1 generation of rats. BMC Complement Altern Med. 17(1):67.

Wang J et. al. (January 2017). Prenatal Exposure to Lipopolysaccharide Alters Renal DNA Methyltransferase Expression in Rat Offspring. PLoS One. 12(1):e0169206.

Wang J et. al. (January 2017). Prenatal Exposure to Lipopolysaccharide Alters Renal DNA Methyltransferase Expression in Rat Offspring. PLoS One. 12(1):e0169206.

Bak ST et. al. (December 2016). Evaluating the Feasibility of DNA Methylation Analyses Using Long-Term Archived Brain Formalin-Fixed Paraffin-Embedded Samples. Mol Neurobiol.

Yi-ping TIAN et. al. (November 2016). Global changes of 5-hydroxymethylcytosine and 5-methylcytosine from normal to tumor tissues are associated with carcinogenesis and prognosis in colorectal cancer JZUSB.

Tomšík P et. al. (September 2016). Boldine Inhibits Mouse Mammary Carcinoma In Vivo and Human MCF-7 Breast Cancer Cells In Vitro. Planta Med.

Zhu H et. al. (April 2016). Race/Ethnicity-Specific Association of Vitamin D and Global DNA Methylation: Cross-Sectional and Interventional Findings. PLoS One. 11(4):e0152849.

Adamu HA et. al. (February 2016). Perinatal exposure to germinated brown rice and its gamma amino-butyric acid-rich extract prevents high fat diet-induced insulin resistance in first generation rat offspring. Food Nutr Res. 60:30209.

Mendonça A. S. et. al. (July 2015). Global methylation and hydroxymethylation patterns of bovine tissues Brazilian College of Animal Reproduction. 12(3)

Wood CE et. al. (April 2015). Latent carcinogenicity of early-life exposure to dichloroacetic acid in mice. Carcinogenesis.

Sadd BM et. al. (April 2015). The genomes of two key bumblebee species with primitive eusocial organization. Genome Biol. 16(1):76.

Miousse IR et. al. (July 2014). Exposure to low-dose (56)fe-ion radiation induces long-term epigenetic alterations in mouse bone marrow hematopoietic progenitor and stem cells. Radiat Res. 182(1):92-101.

Miousse IR et. al. (May 2014). Epigenetic alterations induced by ambient particulate matter in mouse macrophages. Environ Mol Mutagen. 55(5):428-35.

Nzabarushimana E et. al. (February 2014). Long-term epigenetic effects of exposure to low doses of 56Fe in the mouse lung. J Radiat Res.

Weng X et. al. (February 2014). DNA methylation profiling in the thalamus and hippocampus of postnatal malnourished mice, including effects related to long-term potentiation. BMC Neurosci. 15:31.

Zhu X et. al. (January 2014). BCR-ABL1-positive microvesicles transform normal hematopoietic transplants through genomic instability: implications for donor cell leukemia. Leukemia.

Bellizzi D et. al. (December 2013). The control region of mitochondrial DNA shows an unusual CpG and non-CpG methylation pattern. DNA Res. 20(6):537-47.

Riviere G et. al. (December 2013). DNA methylation is crucial for the early development in the Oyster C. gigas. Mar Biotechnol (NY). 15(6):739-53.

Padmanabhan N et. al. (September 2013). Mutation in folate metabolism causes epigenetic instability and transgenerational effects on development. Cell. 155(1):81-93.

Lou J et. al. (August 2013). Role of DNA methylation in cell cycle arrest induced by Cr (VI) in two cell lines. PLoS One. 8(8):e71031.

Customer Reviews

Rating by N*****************@uth.tmc.edu Verified Purchase Reviewed on: Wednesday 04 February, 2015
Application Description
I really enjoy using the fluorometric DNA quantification kit. I find the protocol easy to understand and utilize and simple to complete within 4 hours. I find it gives consistently reliable and repeatable results. The only drawback I can find to this kit is the size of the wells on the plate. I must transfer my final solution to another ELISA plate due to the fact that the well size is incompatible with my fluorescent plate reader. Other than that minor issue, it is a fantastic product!
Epigentek Product Reviews
Related Products

Researchers Also Purchased
MethylFlash Hydroxymethylated DNA Quantification Kit (Fluorometric)
MethylFlash Hydroxymethylated DNA Quantification Kit (Fluorometric)
FitAmp  Blood and Cultured Cell DNA Extraction Kit
FitAmp Blood and Cultured Cell DNA Extraction Kit
EpiQuik DNMT Activity/Inhibition Assay Ultra Kit (Fluorometric)
EpiQuik DNMT Activity/Inhibition Assay Ultra Kit (Fluorometric)
EpiQuik Nuclear Extraction Kit
EpiQuik Nuclear Extraction Kit
Epigentek is an epigenetics company that specializes in epigenetic kits, antibodies, reagents, and services for epigenetic research in DNA methylation, histone modification, and chromatin studies. Terms & Conditions | Privacy Policy | Site Map
Copyright © 2017 Epigentek Group Inc. All rights reserved.