Cyto-Seq
Cyto-Seq
Gene expression cytometry (CytoSeq) is a protocol that enables gene expression profiling of thousands of single cells. In this method, single cells are first randomly deposited into wells. A combinatorial library of beads with specific capture probes are added to each well. After cell lysis, mRNAs hybridize to beads, which are then pooled for reverse transcription, amplification and sequencing. Deep sequencing provides accurate high coverage gene expression profiles of several single cells.
Pros:
- Can readily scale to 10,000s or 100,000s of cells
- Complements and expands the capabilities of fluorescence or mass spectrometry–based cytometry
- Detects any transcribed mRNA without the limitations of antibody availability
- Enables rare cell characterization on small samples with insufficient cells for traditional flow cytometry
- Allows direct analysis of complex samples of heterogeneous cell size and shape
Cons:
- Sequencing depth requires large number of reads (e.g., 200,000 transcripts per cell requires 2 million reads for 10X coverage – 2 billion reads for 1000 cells)
- Single run can be relatively expensive and time-consuming
- Trade-off between depth of sequencing and differential gene expression