"With this method, we are basically able to spot a needle in a haystack - and we can determine if there are five needles in that haystack or if there are 50," he said.
Irudayaraj and his then-graduate research assistant, Kyuwan Lee, who is the first author of the study, adapted common nanotechnology methods to tackle the challenge of pinpointing mRNA splice variants in a living cell. They fabricated gold nanoparticles - more than 1,000 times smaller than the diameter of a human hair - and tagged them with strands of DNA complementary to BRCA1 mRNA splice variants.
When injected into a cell, the nanoparticles attached to either end of mRNA splice variants, forming structures known as dimers - each "like a couple holding hands," Irudayaraj said.
Because dimers give off a unique signal in the presence of light, the researchers could measure the number of dimers by illuminating the cell with a simple light source. The number of dimers corresponded to the number of BRCA1 mRNA splice variants in a cell.
Light behaves differently when it shines on a single gold particle, allowing the researchers to differentiate between dimers and free-floating gold particles.
The researchers used two methods to quantify the dimers: spectroscopy, which measures the way light scatters when it encounters an object, and a colorimetric image on which dimers show as reddish dots while single gold particles appear green.
The technique can quantify mRNA splice variants in a single cell in about 30 minutes.
Irudayaraj is modifying the system to speed up the process so that it can be used in tissue biopsies.
"If we can quantify key mRNA at single cell resolution in a tissue biopsy, that will be very powerful in terms of refining treatment protocols for key diseases," he said.
|Contact: Natalie van Hoose|