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【产品名称】
 MethylFlash甲基化DNA定量试剂盒(Colorimetric)
【订购货号】
 E14147
【产品CAS号码】
 无
【规格/价格】
 
【预计到货时间】
 30 天
【产品详细说明】
【本产品仅供科学研究使用】
【更新日期】 2015-4-1  
MethylFlash Methylated DNA Quantification Kit (Colorimetric)
 
货号:P-1034
 
MethylFlashTM甲基化DNA定量试剂盒(比色法)提供了一套完整的优化缓冲液和试剂,以ELISA-like微孔板形式,通过特异性检测5-甲基胞嘧啶(5-mC)的水平比色法定量检测总的DNA甲基化。与LUMA或LINE-1,Alu,和LTR-based分析方法相比,MethylFlashTM技术能直接地定量检测实际的总DNA甲基化。
 
此款试剂盒尤其适合与MethylFlash羟甲基化DNA定量试剂盒(比色法)配合使用,可同时定量甲基化DNA和羟甲基化DNA
 
 
产品特色
MethylFlashTM甲基化DNA定量试剂盒(比色法)是一款更完美的试剂盒,与之前的 Methylamp™ 总DNA甲基化定量试剂盒相比,其简化的操作程序,提高了结果重复性。它用一种新颖的方法使其背景信号极低,取消了板子干燥和封闭过程。与色谱为基础的方法比如HPLC和质谱相比,此试剂盒提供了一种经济有效的方法精确地检测5-甲基胞嘧啶的水平。
此试剂盒的优势和特色如下:
1.       简单易操作的步骤比色法分析便捷快速,整个流程在4个小时内完成。
2.       新颖的试剂盒组分使背景信号极低,无需封闭板子,使分析变得简单,精确,可靠且一致性好。
3.       高灵敏性,其检测极限可低至0.2ng的甲基化DNA,接受低至50ng的输入基因组DNA。
4.       在指定的浓度范围内,优化的抗体和增强液可高特异性地结合5mC,与未甲基化的胞嘧啶无交叉反应,与羟甲基胞嘧啶无或小至可忽略的交叉反应。
5.       试剂盒包括通用的阳性和阴性对照,适用于定量任何物种里的甲基化DNA。
6.       微孔板形式使分析更具灵活性:手动或高通量分析。
 
原理与程序
MethylFlashTM甲基化DNA定量试剂盒(比色法)包含了总DNA甲基化定量用的所有必需的试剂。在此试剂盒的分析中,DNA被结合到经特殊处理具有DNA高亲和度的微孔板上。用捕获和检测抗体对甲基化的DNA片段进行检测并通过ELISA-like反应和微孔板分光光度计在450nm处的值进行定量。甲基化DNA的量与检测的OD值是成正比的,试剂盒里提供的公式可计算出两种不同的DNA样品的相对甲基化状态或用一个标准曲线对5mC胞嘧啶进行绝对定量。
 
简单,快速,灵活
整个比色分析方法的简单易操作的步骤,便捷快速,能在4个小时内完成。微孔板形式使得分析更灵活,适用于手动或高通量分析。试剂盒提供的通用阳性和阴性对照用于定量任何物种如哺乳动物,植物,真菌,细菌和病毒中的甲基化DNA。
 
高灵敏性和高特异性
新颖的程序和专有的试剂盒组分可精确地高特异性和高灵敏性地定量甲基化DNA。输入DNA检测极限可低至0.2ng的甲基化DNA。
 
 
Product Overview


 

 

 

The MethylFlash™ Methylated DNA Quantification Kit (Colorimetric) is a complete set of optimized buffers and reagents to colorimetrically quantify global DNA methylation by specifically measuring levels of 5-methylcytosine (5-mC) in a microplate-based format.

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

Product Features
The MethylFlash™ Methylated DNA Quantification Kit (Colorimetric) is a further refinement of our previous Methylamp™ 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:

  • Colorimetric 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 0.2 ng of methylated DNA from just 50 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.

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

 

 

 

Product Details



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 (Colorimetric).


Principle & Procedure
The MethylFlash Methylated DNA Quantification Kit (Colorimetric) 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 through an ELISA-like reaction by reading the absorbance in a microplate spectrophotometer at 450 nm. The amount of methylated DNA is proportional to the OD intensity 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 colorimetric 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 colorimetric 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 0.2 ng of methylated DNA.

 

 

 

Product Components


ME1 (10X Wash Buffer)
ME2 (Binding Solution)
ME3 (Negative Control, 20 µg/ml)*
ME4 (Positive Control, 20 µg/ml)*
ME5 (Capture Antibody, 1000 µg/ml*
ME6 (Detection Antibody, 400 µg/ml)*
ME7 (Enhancer Solution)*
ME8 (Developer Solution)
ME9 (Stop Solution)
8-Well Assay Strips (With Frame)
User Guide

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

Note: The ME3 Negative Control is an unmethylated polynucleotide containing 50% of cytosine. The ME4 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


Ortiz, R. et. al. Global DNA Methylation in Peripheral Blood mononuclear cells predicts prolonged elevations of plasma Il-6 in Women throughout the Breast Cancer Trajectory. Abstract

Hsu, A. et. al (October 2011). Promoter de-methylation of Cyclin D2 by sulforaphane in prostate cancer cells. Clinical Epigenetics. 3:3.Full Text Article

Ward, A. et. al. (February 2011). Aberrant methylation of Polo-like kinase CpG islands in Plk4 heterozygous mice. BMC Cancer. 11:71.Full PDF Article

Li, W. et. al. (2011). Distribution of 5-hydroxymethylcytosine in different human tissues. J Nucleic Acids. 2011:870726. PubMed Abstract

 

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