Single-cell level characterization of T cells with dCODE Dextramer® and BD Rhapsody

Multi-Omics Characterization of T-Cell Populations at the Single-cell Level Utilizing Sensitive dCODE Dextramer® and BD® AbSeq on the BD Rhapsody™ Single-Cell Analysis System

Jacobsen, K. et al. Poster presented at SITC 2020

Background

This study aimed to detect and characterize disease-specific CD4+ T cells within thousands of PBMC by combining two technologies: dCODE Dextramer® (Immudex) and single-cell multi-omics analysis using BD Rhapsody™ Single-Cell Analysis System.

dCODE Dextramer®

Workflow Approach

dCODE Dextramer®(RiO) reagents are designed to be compatible with BD’s Rhapsody™ Single Cell Analysis System. Each reagent has a unique DNA barcode specific for the MHC-peptide displayed on the dCODE Dextramer® and the barcode can be amplified and sequenced simultaneously with targeted mRNA from a single cell.

  1. PBMCs stimulated with two peptide antigens (EBV and Tetanus toxoid) were stained with a panel of dCODE Dextramer® (DRB1*0101/EBV, DRB1*0101/TT and DRB1*0101 / DRB1*0101/negative control peptide)
  2. Stained cell sample was captured as single-cells by BD Rhapsody™ Single-Cell Analysis System
  3. Upon cell lysis, polyadenylated mRNA and dCODE Dextramer® sequences from the same cell were captured and cDNA synthesized
  4. mRNA and dCODE DNA libraries were amplified by PCR, sequenced, followed by data analysis using SeqGeq    

Results

EBV-specific T cells were identified in PBMC samples and no specific cells were detected by the negative control Dextramer® (Fig.1). 

 Fig.1 T-SNE plots showing detection of EBV dCODE Dextramer® + cells

 

Frequency of TT-specific T cells was very low, though detectable (Fig.2).
Differential gene expression analysis was seen between Dextramer® EBV+ population vs. EBV-Negative and showed concordance with activated T cells.

 

Fig.2 Detected rare TT Dextramer® + cells (dot-plot)

Conclusions

  • dCODE Dextramer® is compatible with BD Rhapsody™ Single-Cell Analysis System workflow
  • All dCODE Dextramer® used to stain cells were detected in sequencing experiments and antigen-specific T-cell detection results further verified by conventional FACS
  • Distinct cell populations with even low frequencies T cells could be identified in donor sample and their gene expression profile mapped at single-cell level
  • High-resolution T-cell profiling has broad implications in immune-oncology, autoimmunity, and infectious disease

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For research use only. Not for use in diagnostic or therapeutic procedures. Immudex is the sole manufacturer and provider of dCODE Dextramer® (RiO) reagents and support related to these products is through Immudex.