Tissue heterogeneity has pestered researchers desiring a closer look at tissue-specific genomics and epigenomics. Sure, laser capture microdissection (LCM) platforms have been really helpful in isolating very small and specific regions of tissue, but that’s only half the battle.
Working with minute amounts of LCM samples, particularly when they’re derived from the battered formalin-fixed paraffin-embedded (FFPE) samples that comprise a huge portion of the worlds tissue samples, is challenging on the best of days. Moreover, bisulfite-conversion, the reigning sample prep champ for locus-specific methylation analysis, has a reputation for damaging pristine samples, let alone the already roughed up FFPE collections, so researchers looking to profile DNA methylation can be in for a rough road to useable data.
A recently published method developed by researchers at Epigenomics might provide some hope. It uses a combination of laser capture microdissection (LCM) to tease apart the sample, purification and bisulfite treatments optimized for just a few cells, and multiplex and singleplex PCR protocols, to simultaneously compare the methylation status of several genes in individual cell types. A marker incorporated into the PCR reaction allows for normalization of the products during the subsequent quantitative sequencing reaction.
When the team looked at tissue from breast cancer patients, comparing various cell types from normal and malignant cells, they found a difference in methylation status at several loci. Check out the details in J Histochem Cytochem, January 2009.