title: Ultra-sensitive detection of rare T cell clones.
abstract: Advances in high-throughput sequencing have enabled technologies that probe the adaptive immune system with unprecedented depth. We have developed a multiplex PCR method to sequence tens of millions of T cell receptors (TCRs) from a single sample in a few days. A method is presented to test the precision, accuracy, and sensitivity of this assay. T cell clones, each with one fixed productive TCR rearrangement, are doped into complex blood cell samples. TCRs from a total of eleven samples are sequenced, with the doped T cell clones ranging from 10% of the total sample to 0.001% (one cell in 100,000). The assay is able to detect even the rarest clones. The precision of the assay is demonstrated across five orders of magnitude. The accuracy for each clone is within an overall factor of three across the 100,000 fold dynamic range. Additionally, the assay is shown to be highly repeatable.
authors.affiliation: Fred Hutchinson Cancer Research Center, Seattle, WA 98109, United States. hrobins@fhcrc.org
authors.forename: Harlan
authors.initials: H
authors.lastname: Robins
authors.forename: Cindy
authors.initials: C
authors.lastname: Desmarais
authors.forename: Jessica
authors.initials: J
authors.lastname: Matthis
authors.forename: Robert
authors.initials: R
authors.lastname: Livingston
authors.forename: Jessica
authors.initials: J
authors.lastname: Andriesen
authors.forename: Helena
authors.initials: H
authors.lastname: Reijonen
authors.forename: Christopher
authors.initials: C
authors.lastname: Carlson
authors.forename: Gerold
authors.initials: G
authors.lastname: Nepom
authors.forename: Cassian
authors.initials: C
authors.lastname: Yee
authors.forename: Karen
authors.initials: K
authors.lastname: Cerosaletti
mesh_terms: D002999:Clone Cells; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D011948:Receptors, Antigen, T-Cell / Q000235:genetics*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D011948:Receptors, Antigen, T-Cell