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J Immunol. 2015 Jun 15;194(12):6155-63. doi: 10.4049/jimmunol.1500215. Epub 2015 May 8.

Quantitative profiling of immune repertoires for minor lymphocyte counts using unique molecular identifiers.

Author information

1
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia;
2
Vavilov Institute of General Genetics, Russian Academy of Sciences, 119991 Moscow, Russia;
3
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia; Faculty of Medicine, Lomonosov Moscow State University, 119192 Moscow, Russia;
4
Central European Institute of Technology, Masaryk University, 601 77 Brno, Czech Republic; and.
5
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.

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.

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PMID:
25957172
DOI:
10.4049/jimmunol.1500215
[Indexed for MEDLINE]
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