title: Quantitative profiling of immune repertoires for minor lymphocyte counts using unique molecular identifiers.
abstract: Emerging high-throughput sequencing methods for the analyses of complex structure of TCR and BCR repertoires give a powerful impulse to adaptive immunity studies. However, there are still essential technical obstacles for performing a truly quantitative analysis. Specifically, it remains challenging to obtain comprehensive information on the clonal composition of small lymphocyte populations, such as Ag-specific, functional, or tissue-resident cell subsets isolated by sorting, microdissection, or fine needle aspirates. In this study, we report a robust approach based on unique molecular identifiers that allows profiling Ag receptors for several hundred to thousand lymphocytes while preserving qualitative and quantitative information on clonal composition of the sample. We also describe several general features regarding the data analysis with unique molecular identifiers that are critical for accurate counting of starting molecules in high-throughput sequencing applications.
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Evgeny S
authors.initials: ES
authors.lastname: Egorov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Ekaterina M
authors.initials: EM
authors.lastname: Merzlyak
authors.affiliation: Vavilov Institute of General Genetics, Russian Academy of Sciences, 119991 Moscow, Russia;
authors.forename: Andrew A
authors.initials: AA
authors.lastname: Shelenkov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Olga V
authors.initials: OV
authors.lastname: Britanova
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia; Faculty of Medicine, Lomonosov Moscow State University, 119192 Moscow, Russia;
authors.forename: George V
authors.initials: GV
authors.lastname: Sharonov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Dmitriy B
authors.initials: DB
authors.lastname: Staroverov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Dmitriy A
authors.initials: DA
authors.lastname: Bolotin
authors.affiliation: Central European Institute of Technology, Masaryk University, 601 77 Brno, Czech Republic; and.
authors.forename: Alexey N
authors.initials: AN
authors.lastname: Davydov
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Barsova
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
authors.forename: Yuriy B
authors.initials: YB
authors.lastname: Lebedev
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia; Central European Institute of Technology, Masaryk University, 601 77 Brno, Czech Republic; and Pirogov Russian National Research Medical University, 117997 Moscow, Russia ChudakovDM@mail.ru mikhail.shugay@gmail.com.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Shugay
authors.affiliation: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia; Central European Institute of Technology, Masaryk University, 601 77 Brno, Czech Republic; and Pirogov Russian National Research Medical University, 117997 Moscow, Russia ChudakovDM@mail.ru mikhail.shugay@gmail.com.
authors.forename: Dmitriy M
authors.initials: DM
authors.lastname: Chudakov
mesh_terms: D019295:Computational Biology / Q000379:methods; D018076:DNA, Complementary; D020869:Gene Expression Profiling / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D018655:Lymphocyte Count; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D056510:Position-Specific Scoring Matrices; D011947:Receptors, Antigen, B-Cell / Q000737:chemistry / Q000235:genetics; D011948:Receptors, Antigen, T-Cell / Q000737:chemistry / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D011947:Receptors, Antigen, B-Cell; D011948:Receptors, Antigen, T-Cell