Cells communicate through physical interaction of cell surface proteins or through autocrine, paracrine, juxtacrine or endocrine mechanisms. However, cell types identified by current large-scale single-cell sequencing and spatial transcriptomics efforts may mask heterogeneity in functional responses between cell types. Our ultimate vision for a product would be a platform to interrogate multiple cell-types in different combinations and under different biochemical conditions to understand heterogenous gene expression responses. In phase I of this SBIR, Scribe Biosciences will adapt its proprietary Printed Droplet Microfluidics (PDM) technology to study the impact of perturbations on single-cell transcriptomics. PDM can precisely deliver reagents and single cells to a microwell array at any time, enabling flexible multi-step workflows while maintaining high experimental throughputs. PDM will be used to encapsulate single-cells, incubate them either with an agonist/antagonist or additional cell(s) of different cell-types, and perform scRNA-seq to understand induced transcriptional changes. A novel lipid-barcoding scheme will be used to link cells encapsulated in the same droplets, enabling cells to be recovered with traceable signatures in bulk and sequenced by established single- cell methods.
In Specific Aim 1 we will demonstrate the ability to perturb and incubate single cells in a test system where single Jurkat cells isolated in droplets are treated with anti-CD3 and anti-CD28 antibodies, then assayed for IL2p expression.
In Specific Aim 2 we will demonstrate a barcoding method to bioinformatically link cells incubated within the same droplet while allowing them to be sequenced separately. The successful execution of this phase I program will demonstrate the technical feasibility of a high throughput functional genomics platform using our PDM technology and will set the stage for a numerical scale up of the platform to analyze up to 10,000 single cells.
Although single cell genomics have become widespread, these technologies do not include information the original biological context of an analyzed sample. Next generation technologies will integrate biochemical and cellular contexts, leading to a more complete understanding of the relationship between functional perturbations and the transcriptomic changes they engender. In this project, we will apply Scribe Biosciences? proprietary Printed Droplet Microfluidics technology to deterministically perturb single cells with agents or other single cells, then use a new barcoding technology link perturbation conditions to single cell sequencing data.
