unravel the specificity of the adaptive immune response · cd16+cd56+6.1% double negative t...
TRANSCRIPT
10xgenomics.com/vdj
PRODUCT SHEET Chromium | Single Cell Immune Profiling
Unravel the specificity of the adaptive immune response
Feature Barcode technology for cell surface protein & antigen specificitySimultaneously examine the cellular context of the adaptive immune response and immune repertoires of hundreds to tens of thousands of T and B cells in human or mouse with the Chromium Single Cell Immune Profiling Solution. Now, you can also detect cell surface markers and determine antigen specificity by leveraging Feature Barcode technology. Decipher the mechanisms involved in the antigen-specific immune response by distinguishing antigen specificity of single T cells with DNA-barcoded peptide-MHC (pMHC) multimers. Additionally, you can gain further insight into
cellular phenotype via the combination of gene expression read-outs and cell surface protein expression using antibodies conjugated to barcodes instead of the fluorescent tags used in flow cytometry. The resulting barcode diversity vastly expands the breadth of cell surface markers that can be detected at the same time. Use the Single Cell Immune Profiling Solution with Feature Barcode technology to unveil a new understanding of the adaptive immune response including T-cell receptor (TCR) specificity, all within the landscape of the responding cell population.
Improved Resolution of Cell Types with the Addition of Cell Surface Protein Detection tSNE projections of ~8000 peripheral blood mononuclear cells (PBMCs) output by Cell Ranger. A. PBMCs are grouped together based on digital gene expression information. B. PBMCs were then grouped by cell surface protein expression profiles. The additional information from antibody UMI counts better resolved many canonical cell types compared to gene expression data alone—double negative and memory T cells, CD4+ and CD8+ T cells, and memory and naive CD4+ or CD8+ T cells (populations denoted with asterisk*), for example.
tSNE1
A. Gene Expression
B. AntibodytSNE1
tSN
E2tS
NE2
T Regulatory CD4+CD127-CD25+1.6%
Cytotoxic T Naive CD3+CD8a+CD45RA
+5.2%
Helper T Memory CD3+CD4+CD45RA-19.2%
Monocyte Classical CD14+CD16-31.7%
B cells CD19+13.9%
Natural Killer CD16+CD56+6.1%
Double Negative T CD3+CD4-CD8a-3.0%
Cytotoxic T Memory CD3+CD8a+CD45RA-5.5%
Helper T Naive CD3+CD4+CD45RA+9.6% Monocyte Non-classical CD14+CD16+1.5%
T Regulatory CD4+CD127-CD25+1.6%
Helper T Memory CD3+CD4+CD45RA-19.2%
Monocyte Classical CD14+CD16-31.7%
B cells CD19+13.9%
Natural Killer CD16+CD56+6.1%
Helper T Naive CD3+CD4+CD45RA+9.6%
Monocyte Non-classical CD14+CD16+1.5%
Cytotoxic T Memory CD3+CD8a+
CD45RA-5.5%*
Double Negative T CD3+CD4-CD8a-3.0%*
Cytotoxic T Naive CD3+CD8a+
CD45RA+5.2%*
*
10xgenomics.com/vdj
Unravel the specificity of the adaptive immune response
2
Highlights• Reveal clonality, diversity, antigen
specificity, and cellular context
• Pair α and β chain TCR sequences from individual T cells
• Pair heavy and light chain immunoglobulin (Ig) sequences from individual B cells with full isotype resolution
• Simultaneously measure TCR, B cell Ig, cell surface protein expression, and 5’ gene expression in the same cells
• Link full-length, paired TCR α and β chain sequences with TCR-pMHC specificity
Solution Features• Ready-to-use, robust protocols
including Feature Barcode technology
• Compatible partners for oligo conjugated antibodies and MHC multimers
• Documentation for custom conjugations for use with Feature Barcode technology
• Easy to use software includes Cell Ranger Analysis Pipelines, Loupe Browser and Loupe V(D)J Browser visualization tools
System Features• Partition 100 – 80,000+ cells
in under 7 minutes
• Superior sensitivity
• Simple workflow
• Cell size flexibility, no lower limits
• High cell capture rates of up to 65%
• Low doublet rates of under 0.9% in 1000 cells
Product sheet
10 0 10 1 10 2 10 3 10 4 10 5
0
50
100
150
PE Positive75.5
10 0 10 1 10 2 10 3 10 4 10 5
0
20
40
60
80
100
PE Positive35.2
Flow
Cyt
omet
ry
(Cou
nts)
CMV EBV
Feat
ure
Barc
ode
Tech
nolo
gy
(Cou
nts)
Detection of pMHC Specificity in Antigen-Expanded Cells Commercially available CD8+ T cells expanded against CMV and EBV antigens (CMV: HLA-A*0201 NLVPMVATV, EBV: HLA-A*0201 GLCTLVAML, Astarte Biologics). Flow cytometry and Feature Barcode technology identify similar Dextramer-binding cell populations when profiled with dCODE Dextramer reagents displaying the same pMHC specificities.
10xgenomics.com/spatial-gene-expression3
Unravel the specificity of the adaptive immune response Product sheet
V D J C
α TRAV26-2 - TRAJ43 TRAC
α TRAV40 - TRAJ53 TRAC
β TRBV30 - TRBJ2-4 TRBC2
CD3
CD19
CMV Dextramer EBV Dextramer NC Dextramer
CD4
CD45RA
CD8a
CD45RO
TCRα: TRAV26-2 : TRAJ43 : TRACTTTTATGGGGA A A ATTGA A ACCTGCCTGATGTGGGATGTGCTGTGGCTGCTGCTTTGTTGCTTGGGACCTCCTCTGACCTA GGATCAGACACAGAGTCTGAGTTCTGGGGCCTGGA ACCTCA ATGTGCACTTGA ACA ATGA AGTTGGTGACA AGCATTACTG TACTCCTATCTTTGGGTATTATGGGTGATGCTAAGACCACACAGCCAAATTCAATGGAGAGTAACGAAGAAGAGCCTGTTCACTTG CCTTGTAACCACTCCACAATCAGTGGAACTGATTACATACATTGGTATCGACAGCTTCCCTCCCAGGGTCCAGAGTACGTGAT TCATGGTCTTACA AGCA ATGTGA ACA ACAGA ATGGCCTCTCTGGCA ATCGCTGA AGACAGA A AGTCCAGTACCTTGATCCT GCACCGTGCTACCT TGAGAGATGCTGCTGTGTACTACTGCATCCTGA ACA ATA ACA ATGACATGCGCT T TGGAGCAGGGA CCAGACTGACAGTAAAACCAAATATCCAGAACCCTGACCCTGCCGTGTACCAGCTGAGAGACTCTAAATCCAGTGACAAGTC TGTCTGCCTATTCACCGATTTTGATTCTCAAACAAATGTGTCACAAAGTAAGGATTCTGATGTGTATATCACAGACAAAACTGT GCTAGACATGAGGTCTATGGACTTCAAGAGCAACAGTGCTGTGGCCTGGAGCAACAAATCTGACTTTGCATGTGCAAACGCCTTCA
TCRα: TRAV40 : TRAJ53 : TRACG A AT G AG G AC AG TAG T G AG G AC AC AC T G C AG A AC AC T G AG AG G C AG AT G AC G T C T TATA AC C T G T T C AT TA AC T T G C C A C T T G G G G G T G C TAT T C A C C AG T G A A AT C ATA AT T TAT G C AT G A AT T C T G AG TAC A AC T G T G A AT C C T C A C T T C A ACAGTG ATG CCCTCTG CTAGG CCAG AG ACACTA ACA ATG A ACTCCTCTCTGG ACT T TCTA AT TCTG ATCT TA ATGT T TG GAGGAACCAGCAGCAATTCAGTCAAGCAGACGGGCCAAATAACCGTCTCGGAGGGAGCATCTGTGACTATGAACTGCACAT ACACATCCACGGGGTACCCTACCCTTTTCTGGTATGTGGA ATACCCCAGCA A ACCTCTGCAGCTTCTTCAGAGAGAGACA A TGG A A A ACAG CA A A A AC T TCGG AGG CGG A A ATAT TA A AG ACA A A A AC TCCCCCAT TGTG A A ATAT TCAGTCCAGGTA TCAGACTCAGCCGTGTACTACTGTCT TCCGCTGCA AGGAGGTAGCA ACTATA A ACTGACAT T TGGA A A AGGA ACTCTCT T A ACCGTGA ATCCA A ATATCCAGA AGCCTGACCCTGCCGTGTACCAGCTGAGAGACTCTA A ATCCAGTGACA AGTCTGTCT GCCTATTCACCGATTTTGATTCTCA A ACA A ATGTGTCACA A AGTA AGGATTCTGATGTGTATATCACAGACA A A ACTGTGCTA GACATGAGGTCTATGGACTTCAAGAGCAACAGTGCTGTGGCCTGGAGCAACAAATCTGACTTTGCATGTGCAAACGCCTTCA
TCRβ : TRBV30 : TRBJ2-4 : TRBC2TTATGGGGGCAGAGTATAGCTGGTTCCCAGGGAGGCTGGGGGTGATTCACCACACTCTTAAAAGAAGACTAGCCTCATTTCCT CCCCAGCTCATGGGGTAGGATTCCTCTGCTCTGGCAGCAGATCTCCCAGAGGGAGCAGCCTGACCACATCACTGGCCCAGA AGAGGAGGCGTCTGTCCCCCAGACTAGCTGAAGGAAAGGCTGGCTTGGATGATGCTCTGCTCTCTCCTTGCCCTTCTCCTGG GCACTTTCTTTGGGGTCAGATCTCAGACTATTCATCAATGGCCAGCGACCCTGGTGCAGCCTGTGGGCAGCCCGCTCTCTCTG GAGTGCACTGTGGAGGGAACATCAAACCCCAACCTATACTGGTACCGACAGGCTGCAGGCAGGGGCCTCCAGCTGCTCTTCTACT CCGTTGGTATTGGCCAGATCAGCTCTGAGGTGCCCCAGAATCTCTCAGCCTCCAGACCCCAGGACCGGCAGTTCATCCTGAGTTCT A AGA AGCTCCTTCTCAGTGACTCTGGCTTCTATCTCTGTGCCTGGAGTATCTCAGACCTAGCCA A A A ACATTCAGTACTTCG GCGCCGGGACCCGGCTCTCAGTGCTGGAGGACCTGAAAAACGTGTTCCCACCCGAGGTCGCTGTGTTTGAGCCATCAGAAGC AGAGATCTCCCACACCCAAAAGGCCACACTGGTGTGCCTGGCCACAGGCTTCTACCCCGACCACGTGGAGCTGAGCTGGTGGGTGAAT GGGAAGGAGGTGCACAGTGGGGTCAGCACAGACCCGCAGCCCCTCAAGGAGCAGCCCGCCCTCAATGACTCCAGATACTGCCTGA
A.
B.
Profile PBMCs with CMV+ T cells Labeled with TotalSeq-C Antibodies and dCODE Dextramers CMV-PBMCs mixed with 2% CMV expanded T cells. A. Stained with a panel of TotalSeq-C antibodies and a panel of 3 dCODE Dextramer reagents displaying CMV, EBV, and negative control (NC) peptides, respectively. Cell populations are clustered based on surface protein expression: T cells (CD3+, CD4+ and CD8+, panels 1-3), and B cells (CD19+, panel 4). CD8+ T cells bind specifically to Dextramers displaying a CMV antigen (panel 7). The sample does not contain EBV-specific T cells, so the EBV Dextramer staining is similar to the negative control Dextramer staining (panels 8-9). B. Paired T cell receptor clonotypes are assembled for the CMV-specific T cells; these CMV+CD8+ T cells are highly expanded and experienced (CD45RO+). The table outlines gene calls for the most prevalent TCR clonotype alpha (α) and beta (β) chains. The paired α and β chain V(D)J sequences are shown and corresponding V(D)J nucleotides are color-coded (5’UTR: gray, V: red, D: yellow, J: green, C: purple).
© 2020 10x Genomics, Inc. FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.LIT000033 Rev D Chromium Single Cell Immune Profiling Feature Barcode Technology Product Sheet
Products Product Code
Chromium Single Cell 5’ Library Construction Kit, 16 rxn 1000020
Chromium Single Cell 5’ Library & Gel Bead Kit, 16 rxn1 1000006
Chromium Single Cell 5’ Library & Gel Bead Kit, 4 rxn1 1000014
Chromium Single Cell V(D)J Enrichment Kit, Human T Cell, 96 rxn 1000005
Chromium Single Cell V(D)J Enrichment Kit, Human B Cell, 96 rxn 1000016
Chromium Single Cell V(D)J Enrichment Kit, Mouse T Cell, 96 rxn 1000071
Chromium Single Cell V(D)J Enrichment Kit, Mouse B Cell, 96 rxn 1000072
Chromium i7 Multiplex Kit, 96 rxn 120262
Chromium i7 Multiplex Kit N, Set A, 96 rxn 1000084
Chromium Single Cell A Chip Kit, 16 rxn1
1000009 (EMEA) 1000151(US & APAC)
Chromium Single Cell A Chip Kit, 48 rxn1
120236 (EMEA) 1000152 (US & APAC)
Chromium Single Cell 5’ Feature Barcode Library Kit, 16 rxn 1000080
Chromium Controller & Accessory Kit, 12 Mo. Warranty 120223
Chromium Controller & Accessory Kit, 24 Mo. Warranty 120246
Cell Ranger go.10xgenomics.com/vdj/cell-ranger
Loupe Browser go.10xgenomics.com/vdj/loupe-cell
Loupe V(D)J Browser go.10xgenomics.com/vdj/loupe-vdj
Compatible Partner Product: Biolegend TotalSeq™-C
go.10xgenomics.com/totalseq-C
Compatible Partner Product: Immudex dCODE™ Dextramers®
go.10xgenomics.com/dCODE-Dextramers
Research areas• Basic & Translational Immunology
• Immuno-oncology & Immunotherapy
• Autoimmune Disorders & Inflammatory Diseases
• Infectious Disease & Vaccine Research
• Transplant & Immune Reconstitution
Applications • Immune Profiling
• T Cell Antigen Specificity
• Tumor Infiltrating Lymphocyte Characterization
• Immune Derived Cellular Therapy Discovery
• Characterization of the Tumor Microenvironment
• Immune Cell Atlasing
Additional resourcesDatasets go.10xgenomics.com/vdj/datasets
Seminars go.10xgenomics.com/vdj/seminars
Application Notes go.10xgenomics.com/vdj/app-notes
Technical Support go.10xgenomics.com/vdj/support
Publications go.10xgenomics.com/vdj/pubs
Unravel the specificity of the adaptive immune response Product sheet
1 Available for use only with the indicated Chromium Controller (PN-120223;120246) or Chromium Single Cell Controller (PN-120263;120212).
Chromium Chip A Single Cell Kit product code is region specific and should be used based on customer’s geographical location.
US: America; APAC: Asia Pacific; EMEA: Europe, Middle East and Africa