The application of chromatin immunoprecipitation (ChIP) in epigenetics research is expanding constantly. Lately it seems that every last aspect of the ChIP process from antibody validation and DNA shearing to purification is being fine-tuned by experts worldwide.
One of the latest upgrades to the ChIP process improves chromatin capture and cleanup. Over the years researchers have deployed sepharose columns, agarose beads, or more recently, magnetic beads to separate the wheat from the chaff in immunoprecipitations. Scientists at Porvair Filtration set out to improve upon this key ChIP step with a new technology using BioVyon solid phase, porous polymer discs mounted in spin columns.
In a published evaluation, (Chernukhin et al, Analytical Biochemistry, May 2011 scientists from the University of Essex pitted the BioVyon affinity matrix developed by Porvair Filtration Group, against other established ChIP enrichment methods.
The Essex team compared the new BioVyon matrix to other common techniques (Protein A Sepharose, and Dynabeads Protein A) in ChIP assays using two antibodies and seven DNA loci, and found that it outperformed the bead-based approaches. When the dust settled, the researchers uncovered some interesting benefits of the new approach.
BioVyon Protein A had better percentage of DNA pull-down in all of the assays.
- Signal to noise ratios were improved with Biovyon. 4-fold enrichments were seen in 20/21 assays vs. 16 for Dynabeads and 13 for Sepharose.
- There was less processing using the solid phase approach. The IPs took 10-15 minutes, had less washing and no separation steps, which minimized sample loss and handling errors for the scientists.
- The authors thought that these attributes would help in small volume micro-ChIP or automated high throughput ChIP assays.
Polymer Discs Pay Off in ChIP
Porvair has made the BioVyon discs commercially available in their new Chromatrap® kits. Despite being a very recent innovation, some researchers are already putting the columns to work, like Dr Ronald N Hines, Professor of Pediatrics, Pharmacology & Toxicology at the CRI. “I believe the bead-based system gave us a relatively small signal to noise ratio such that even minor variation deviations caused by variation in the efficiency of the bead system (for whatever reason) resulted in considerable variation in the assay, “shared Hines. “By comparison I think the most substantial advantage of the Chromatrap system is more efficient and less variable trapping of the immunoprecipitated chromatin such that the signal to noise ratio is larger and influenced less by minor variations in trapping efficiency”.
A Close Look at Solid Phase
Kevin Quinlan, Epigenetics Business Manager from Porvair Filtration Group provides an inside look at how Chromatrap® works. “The chemically inert porous material that forms the support for the anchored capture chemistry has a large internal surface area. The reactions, mixing and washing all take place inside this small controlled volume that enables highly efficient and specific chromatin capture with low background binding.” Plus, as Quinlan explains “Because microbeads are no longer required for performing ChIP assays, Chromatrap® simplifies sample handling and reduces errors associated with pipetting.”
The Chromatrap® crew at Porvair says their columns routinely show:
- DNA pull-down up to 25 times higher than with bead based procedures.
- DNA enrichment with signal to noise ratios 2 to 3 times better.
- Great results with small sample sizes; as few as 15,000 cells per assay.
So, if your beads aren’t cutting it or you want to further streamline your ChIP process, you can learn more about the Chromatrap® System at Porvair’s attractive microsite.