SpeedRun One-Hour MBD3 ELISA Kit (Colorimetric)

The SpeedRun™ One-Hour MBD3 ELISA Kit (Colorimetric) is a complete set of optimized reagents designed for measuring the amount of MBD3 using nuclear extracts or purified MBD3 proteins from a broad range of species such as human, mouse and rabbit, in a variety of forms including, but not limited to, cultured cells and fresh/frozen tissues. The kit has the following advantages and features:

• Quick: Colorimetric assay completed within 1 hour.
• Versatile: Works with nuclear extracts from cells/tissues or purified MBD3 proteins.
• Sensitive: Limit of detection can be as low as 1 ng of MBD3 proteins.
• Convenient: Includes MBD3 protein standard for quantification.
• Flexible: Allows manual or high-throughput analysis with strip microplate format.

Background Information
MBD3 (methyl-CpG-binding domain protein 3) is a member of the MBD protein family. MBD3 is not capable of binding to methylated DNA but instead binds to hydroxymethylated DNA. MBD3 is a subunit of the NURD, a multisubunit complex containing nucleosome remodeling and histone deacetylase activities. MBD3 mediates the association of metastasis-associated protein 2 (MTA2) with the core histone deacetylase complex, linking it to transcriptional control within pathways related to gene expression, chromatin organization, developmental biology, and signal transduction. The binding of MBD3 to hydroxymethylated DNA may be affected by MBD3 mutation and biochemical or pharmacological intervention. Pathogenic variation in MBD3 is reported in autism spectrum disorder. This places the gene in a clinically relevant regulatory network in which altered chromatin control has been associated with neurodevelopmental disease. Thus, detecting level of MBD3 is important in elucidating mechanisms of epigenetic regulation of gene activation and silencing, as well as benefiting cancer and neuro disease therapeutics.

Principle & Procedure
The kit contains all reagents necessary for the quantification of MBD3 levels. In this assay, MBD3 in the extracts is bound to strip-wells with a high binding buffer and is detected using MBD3 Detection Complex and then quantified colorimetrically by reading the absorbance in a microplate spectrophotometer. The MBD3 level is proportional to the OD intensity measured.

Starting Materials
Input materials can be nuclear extracts or purified MBD3 proteins. The amount of nuclear extracts for each assay can be 1 µg to 2 µg. The amount of purified proteins can be 1 ng to 50 ng with an optimal range of 10-20 ng, depending on the purity of the proteins.