page_title: CITE-Seq
section_title: Wet lab workflow
hierarchical_section_title: CITE-Seq / Workflow / Wet lab workflow
caption_attribution_description: English: Schematic of the dry lab workflow for CITE-Seq
context_page_description: CITE-Seq is a method for performing RNA sequencing along with gaining quantitative and qualitative information on surface proteins with available antibodies on a single cell level. So far, the method has been demonstrated to work with only a few proteins per cell. As such, it provides an additional layer of information for the same cell by combining both proteomics and transcriptomics data. For phenotyping, this method has been shown to be as accurate as flow cytometry by the groups that developed it. It is currently one of the main methods, along with REAP-Seq, to evaluate both gene expression and protein levels simultaneously in different species.
  The method was established by the New York Genome Center in collaboration with the Satija lab.
context_section_description: The first step involves preparation of the antibody-oligo conjugates also known as Antibody-Derived Tags (ADTs). ADT preparation involves labeling an antibody directed against a cell surface protein of interest with oligonucleotides for barcoding the antibody.
  Once you have the ADTs, the next step is to bind the cells with the desired ADT pool. The scRNA-seq libraries can be prepared using Drop-seq, 10X Genomics or ddSeq methods. In brief, ADT labelled cells are encapsulated within a droplet as single cells with DNA-barcoded microbeads.
  Within a droplet, the cells are next lysed to release both bound ADTs as well as mRNA. These then are converted to cDNA. Each DNA sequence on a microbead has a unique barcode thus indexing cDNA with cell barcodes.  cDNA is prepared from both ADTs and cellular mRNAs.
  In the next step, based on the developer's guidelines, cDNA is PCR-amplified and ADT cDNA and mRNA cDNA are separated based on size (generally, ADT-derived cDNAs are < 180bp and mRNA-derived cDNAs are > 300bp). Each of the separated cDNA molecules is independently amplified and purified to prepare sequencing libraries. Finally, the independent libraries are pooled together and sequenced. Thus, proteomics and transcriptomics data can be obtained from a single sequencing run.
caption: Schematic of the dry lab workflow for CITE-Seq
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