Single Cell Analysis

While all the cells that comprise an organism may share identical genes, the same cannot be said for each cell’s phenotype. Humans and other eukaryotes have evolved a collection of diverse tissues and specialized cell types, each carrying out distinct functions. Even among similar cell types, evidence continues to suggest that gene expression at this level can also be heterogeneous. To account for the variability that occurs from one closely related cell to another, new methods in single-cell analysis (SCA) have been developed.

What are Applications in Single Cell Analysis?

Single cell applications uncover the hidden nuances that lie within complex cellular populations by simultaneously characterizing each cell along with their phenotypic signatures. Single-cell transcriptomic approaches, such as single-cell RNA sequencing (scRNA-seq), analyze gene transcripts to discover rare cell populations, regulatory relationships, and distinct cell lineages. Downstream at the protein level, proteomic approaches such as SCoPE-MS and nanoPOTS are ideal for mapping cellular proteins in very heterologous tissues (such as tumors) and addressing what may be missed at the genetic level (such as cell-surface proteins and post-translational modifications). Investigations of regulomes and genome-wide regulatory landscapes in single cells are also possible with methods like scATAC-seq and scDNase-seq. Check out the following tools and equipment that can help drive your single cell workflow.

Cell Isolation Tools

As the first step to any single-cell analysis, individual cells must be separated from a more complex population within a liquid suspension or a larger tissue in a manner that is gentle yet efficient. It is ideal to quickly produce purified single cells in a way that minimizes cellular stress or damage as these can affect any underlying structure, function, or phenotype. Scalability for future needs is also a useful consideration. For initiating a single-cell workflow, cell sorters are essential instruments, utilizing technologies such as flow cytometry, fluorescence-based sorting, and specialized microfluidics. These are high-throughput systems that can quickly sort or dispense hundreds to thousands of individual cells. For histological approaches, laser capture microdissection instruments can isolate single cells of interest from microscopic tissue samples without causing cell damage. Finally, our cell isolation catalog lets researchers search for various cell isolation kits, reagents, and accessories, including those designated for particular cell types or isolation methods.

Preparation for Single Cell Sequencing

Isolated samples will need to undergo some preparatory steps before sequencing. Among these are cell lysis, reverse transcription, cDNA synthesis, amplification, library preparation, barcoding, and quality control. Steps in this preparatory workflow will depend on the nature of the sequencing analysis and the desired sequencing platform. For instance, scRNA-seq protocols may require additional cleanup or enrichment to select for the desired mRNA molecule types. To help facilitate these single cell sequencing applications, a variety of sequencing prep kits are available, containing the required sets of reagents and supported by optimized protocols. Prior to sequencing, nucleic acid analysis instruments can be used to for quality control procedures, such as DNA fragment sizing and quantitation. Finally, the limitations and complexity of any sequencing project will depend on the choice of the NGS sequencing platform. The target number of sequenced cells, sequencing depth, read lenght, and sample throughput are some important considerations when planning a sequencing workflow or deciding on a platform.

Specialized Tools

Specialized instruments can benefit SCA workflows by enabling important measurements, automation, and improving throughput. For instance, a variety of cell analysis systems allow analytical modalities, such as automated imaging, cell-based assays, and fluorescent immunophenotyping. Some can also facilitate cell capture, lysis, and library preparation, effectively simplifying the single cell sequencing workflow. Liquid handlers and workstations automate sample preparation procedures, increasing productivity while minimizing human error. Specialized services for single cell-based analysis are also available if your research team lacks the necessary expertise and instrumentation, or if resources need to be allocated to other projects.