How does CITE-Seq work?

CITE-seq combines two processes – single-cell RNA sequencing and cell-surface protein detection – into a single process. The aim is to examine the transcriptome of individual cells while simultaneously also examining the surface proteins expressed on the cell membrane. This gives researchers a better understanding of the functionality and diversity of individual cells in a larger population. 

These are the key steps of CITE-SEQ: 

• Tissue is first dissociated into a single cell suspension and the cells are stained with antibody-oligonucleotide conjugates targeting specific surface proteins of interest. These antibodies are conjugated with unique DNA barcodes instead of fluorescent labels.
• The labeled cells are then encapsulated in small droplets, where each droplet contains a single cell along with reagents for reverse transcription. 
• The reverse transcriptase adds a unique barcode to the mRNA transcripts of the cell and to the antibody-oligonucleotide conjugate to facilitate identification during downstream analysis. 
• Standard techniques are used to sequence RNA and protein from each cell individually and the sequencing data is analyzed to identify individual cells based on their protein markers and gene expression. This provides a comprehensive view of both transcriptome and proteome at the single-cell level.