SBIR-STTR Award

Cell Microsystems is a start-up biotechnology company whose mission is to commercialize a cost effective platform for isolating and manipulating single, viable cells. The company's CellRaft(tm) technology is based on a unique cell array recently developed at the University of North Carolina (UNC) at Chapel Hill, and represents an ideal opportunity for the translation of an academic technology to the marketplace through the NIH sponsored SBIR partnership. In recognition of the growing market of single-cell genetic analysis, we will explore the feasibility of using the CellRaft technology as a sample handling tool for the analysis of single cells by polymerase chain reaction (PCR). The CellRaft(tm) technology is unique in enabling individual cells to be identified by a variety of selection critera followed by isolation with extremely high viability, purity, and yield. In this Phase I SBIR proposal, we will optimize the CellRaft(tm) Array for single-cell isolation towards the applicationof PCR, and optimize the collection and transfer method to deliver single cells from the array to a vessel for conducting PCR. Our academic collaborator at UNC will conduct the cell-based experiments to implement the CellRaft(tm) technology for single-cell PCR analysis through the use of a commercial single-cell PCR kit. The product will be specifically developed to be compatible with existing commercial single-cell PCR kits in collaboration with a company who is a world leader in sample handling and assay products. This collaboration will utilize the company's commercial single cell PCR kit. The studies in Phase I will deliver a market-ready CellRaft Array product optimized and specific for single-cell PCR applications. Our goal is to bridge the CellRaft Array technology with single-cell PCR analysis and promote the use of this technology to the large and growing community in the life science market interested in the molecular analysis of single cells.

Public Health Relevance Statement:


Public Health Relevance:
The unique cell sorting/handling technology being developed by Cell Microsystems Inc. will enable genetic analysis of single cells in broad applications in health, forensics, and basic research.

NIH Spending Category:
Biotechnology

Project Terms:
Bacteria; base; Basic Science; Biological; Biological Assay; biological research; Biological Sciences; Biopsy; Biotechnology; cell behavior; Cell Separation; Cells; Characteristics; Collaborations; Collection; Communities; Complex; Core Facility; cost; cost effective; Crime; Cultured Cells; Development; Devices; Disease; disease diagnosis; Drug Industry; Environment; Europe; Forensic Medicine; forest; Funding; genetic analysis; Genomics; Goals; Grant; Health; Heterogeneity; improved; Individual; innovation; Intellectual Property; interest; Knowledge acquisition; Laboratories; Laboratory Research; Letters; Licensing; Link; Marketing; Measurement; Methods; Microscope; microsystems; Mission; Molecular Analysis; North Carolina; novel; Phase; Polymerase Chain Reaction; Population; Preparation; product development; programs; Protocols documentation; public health relevance; Publications; research and development; Research Institute; research study; Sample Size; Sampling; screening; single cell analysis; Small Business Innovation Research Grant; Solid; Sorting - Cell Movement; Sterility; Surface; System; Techniques; Technology; Testing; Tissues; tool; Transcript; Translations; Tube; United States National Institutes of Health; Universities; Virus

Among the commercially available systems for single cell isolation and next generation sequencing (NGS) sample preparation, none are capable of automating both imaging and NGS sample preparation at high throughput. While single cell genomic analysis has improved in sensitivity and throughput, concomitant improvements in detailed phenotypic characterization of cells has not been integrated into automated workflows, nor have they been scaled beyond a few dozen cells per run. Currently, the only means of biomarker- based sorting of cells prior to single cell sequencing is either fluorescence-activated cell sorting (FACS) or imaging cells on a microscope which is separate from an automated sample preparation instrument (e.g. Fluidigm C1, WaferGen ICELL8). For example, 10X Genomics’ technology relies on upfront FACS purification of cell populations, and still cannot connect these relatively superficial phenotypic observations to downstream NGS data. Fluidigm’s C1 system allows imaging of the approximately 800 cells loaded on a microfluidic device on a separate microscope, but this method results in significant rates of multiplets and requires investigators to construct their own imaging methods, equipment and software, independent of the C1 system. Fluidigm’s Polaris allows both imaging and NGS sample preparation in one instrument but is limited in throughput to 48 cells per run and represents a significant capital expense. To address the unmet need for integrated imaging and NGS sample preparation in a single, high-throughput, cost-effective system, Cell Microsystems proposes here the development of the AIR-FLOW™ System. Using our core CellRaft Technology, the AIR-FLOW™ will allow multi-channel fluorescent imaging, isolation of single cells and RNA-Seq library preparation. The AIR-FLOW™ System will allow multi-channel automated imaging of surface markers, morphology and even subcellular features, while integrating a microfluidic sample preparation method developed by Peter Sims, PhD of Columbia University to construct next generation sequencing (NGS) transcriptomic libraries. Dr. Sims’ sample preparation technology is highly complementary with the CellRaft technology: both rely on microwell arrays, fluorescence imaging and the same biocompatible materials. This approach employs optically barcoded beads for mRNA capture and sample preparation, allowing the resulting sequencing data to be directly linked to imaging data on a cell-to-cell basis. Also, by integrating these two technologies, throughputs of several thousand cells per 4-5 hr run will be easily achievable. Based on our Phase I data, Cell Microsystems’ core CellRaft Technology, provides key advantages over existing microfluidic technologies, effectively eliminating cell-to-cell cross-contamination, reducing sample input requirements and providing a less stressful environment which reduces transcriptomic artifacts. During this Phase II program, we will integrate Dr. Sims’ technology with the core CellRaft Technology in the AIR-FLOW™ System.

Public Health Relevance Statement:
Project Narrative Single cell sequencing is often accomplished by fluorescence-based sorting of cells to purify a population of interest, followed by droplet- or microfluidics-based preparation of a sequencing library. After sequencing however, phenotypic imaging and fluorescent marker data is left unconnected to genomic and/or transcriptomic analysis on a cell-to-cell basis. A new system, the Cell Microsystems’ AIR-FLOW™ is proposed here, and will allow imaging-based sorting of single cells followed by integrated microfluidic transcriptomic sample preparation. Among the commercially available options, no single instrument is capable of this fully-integrated workflow.

Project Terms:
Address; aqueous; base; Biocompatible Materials; Biological Markers; Capital; Cell Separation; Cells; cellular imaging; Communities; Complement; Complement component C1s; Computer software; cost effective; Cytolysis; Data; design; Detection; Development; Doctor of Philosophy; Ensure; Environment; Equipment; experimental study; Feedback; Fluorescence; fluorescence imaging; Fluorescence-Activated Cell Sorting; fluorophore; Genomics; Image; image reconstruction; imaging modality; improved; Individual; instrument; interest; Label; Laboratories; Left; Libraries; Link; Liquid substance; Messenger RNA; Methods; Microfluidic Microchips; Microfluidics; Microscope; microsystems; Molecular Biology; Morphologic artifacts; Morphology; multiplet; Needles; next generation sequencing; Oils; oligo (dT); Oligonucleotides; Optics; Performance; Phase; Phenotype; Population; Preparation; programs; Protocols documentation; prototype; Reaction; Reagent; Research Personnel; Running; Sampling; seal; single cell sequencing; Solid; Sorting - Cell Movement; Surface; System; Technology; Temperature; Testing; transcriptome sequencing; transcriptomics; Universities; Validation; Virginia