ChIP has been a dear friend to researchers studying histone modifications for years. But, just like most friends, it has its imperfections too, particularly when it comes to sensitivity. On its best day ChIP is about as sensitive as Mr. Trump and just like The Donald, it can be temperamental. So we were excited to see a team of researchers who have developed a way to zoom into specific cell types in tissue to map out the histone modifications at specific loci.
The method, called in situ hybridization-proximity ligation assay (ISH-PLA), is, you guessed it, a combination of in situ hybridization and proximity ligation.
The researchers wanted a way to get around some of the sensitivity/specificity shortcomings of the ChIP assay and here’s how they tackled it:
- First, ISH. Probes with biotin residues on them anneal to the gene region of interest in tissue slices.
- Then, PLA. Primary antibodies against biotin and against the histone modification you’re interested in bind to biotin and the histone modification.
- Next, secondary antibodies with a PLA probe bind to the primary antibodies.
- Finally, you do the ligation and amplification.
Using this approach, the team was able to:
- Map histone modifications at a given gene locus
- In single cells in vivo, in fixed histological specimens.
“This methodology has promise for broad applications in the study of epigenetic mechanisms in complex multicellular tissues in development and disease,” they conclude.
As you can imagine, with any staining technique comes a ton of pretty data images that you can check out for yourself along with their model data at: Nature Methods, January 2013.