371525 Ultrasensitive Microfluidic ChIP-Seq with Small Number of Cells
Chromatin immunoprecipitation (ChIP) assay has become the technique of choice for examining endogenous DNA-protein interactions over the years. However, conventional ChIP requires a large number of cells (~106~107 cells), which hinders its application to scarce primary samples from animals and patients.
In this project, we developed an ultrasensitive microfluidic ChIP (chromatin immunoprecipitation) assay followed by next generation sequencing technology. The prototypical microfluidic chip, which was fabricated by multilayer soft lithography, is able to manipulate magnetic IP beads in various ways, including formation of compact packed bed (for surface adsorption of target molecules), oscillatory movement (for removal of nonspecific adsorption), and retention of beads by a magnetic field. Thus, all steps involved in immunoprecipitation can be handled by our microfluidic chip without manual operation. Taking advantage of tiny reaction volume, our microfluidic technology leads to dramatic improvement over the current state-of-the-art on both sensitivity and efficiency, therefore it can greatly benefit epigenetic studies of rare or scarce primary cell samples, such as stem cells. As for preliminary results, we were able to produce high-quality ChIP DNA of 1~2 ng from 10,000 cells and 0.2~0.5ng from 3000 cells . Such ChIP efficiency was roughly over 100 higher than that of current state-of-the-art (~10 pg from 10,000 cells). By using the ChIPed DNA, we were able to achieve whole genome mapping of target protein (H3k4me3) binding with as few as 1000 cells. Moreover, the automated microfluidic based platform can dramatically reduce the total time (from ~3 days to several hours) and the hands-on time required for conventional ChIP assay and has great potential for future scale-up.