Single-Cell Multiomics Protocols
Sample Preparation and Cell Labeling
Preparing Single Cell Suspensions
This protocol describes preparation of single cell suspensions of commonly used immune cell types. A single cell suspension can be loaded into a BD Rhapsody™ Cartridge. This protocol does not describe tissue dissociation.
Co-staining cells with fluorescent antibodies and BD Oligo-Conjugated Antibodies
In this technical bulletin, we address how to co-stain cells with the same clone of both fluorescent antibodies and oligonucleotide-conjugated antibodies (BD® Single-Cell Multiplexing Kit or BD® AbSeq Ab-Oligos).
Single Cell Labelling with the BD® Single-Cell Multiplexing Kits
The BD® Human Single-Cell Multiplexing Kit (Cat. No. 633781) and the BD® Mouse Immune Single-Cell Multiplexing Kit (Cat. No. 633793) utilize an innovative antibody-oligo technology to provide higher sample throughput for single cell library preparation.
Single Cell Labeling with BD® Flex Single-Cell Multiplexing Kits
BD® Flex Single-Cell Multiplexing Kits utilize an innovative antibody-oligo technology to provide higher sample throughput for single-cell library preparation, as well as improve detection and elimination of multiplet data from single-cell experiments.
Single Cell Labelling with BD® AbSeq Ab-Oligos (1 plex to 40 plex)
This protocol describes use of BD AbSeq Ab-Oligos (antibody-oligonucleotides) for antigen-expression profiling with BD Rhapsody™ single cell capture and downstream library preparation.
Single-Cell Labeling with BD® AbSeq Ab-Oligos (41 plex to 100 plex)
This protocol describes the use of BD® AbSeq Ab-Oligos (antibody-oligonucleotides) for antigen expression profiling with BD Rhapsody™ single-cell capture and downstream library preparation.
dCODE Dextramer® (RiO) staining protocol
This protocol provides instructions on cell labeling with dCODE Dextramer® and optional BD AbSeq Ab-Oligos.
Single Cell Labelling with the BD® Single-Cell Multiplexing Kit and BD® AbSeq Ab-Oligos (1 plex to 40 plex)
This protocol describes use of BD® AbSeq Ab-Oligos (antibody-oligonucleotides) with the BD® Human Single-Cell Multiplexing Kit (Cat. No. 633781) or BD® Mouse Immune Single-Cell Multiplexing Kit (Cat. No. 633793).
Single-Cell Labeling with BD® Single-Cell Multiplexing Kit and BD® AbSeq Ab-Oligos (41 plex to 100 plex)
This protocol describes the use of BD AbSeq Ab-Oligos (antibody-oligonucleotides) (41 plex to 100 plex) with the BD® Human Single-Cell Multiplexing Kit (Cat. No. 633781).
Single-Cell Labeling with BD® AbSeq Ab-Oligos for Intracellular CITE-seq
This protocol details how to stain for intracellular antigens using BD® Intracellular AbSeq (IC-AbSeq) antibodies for profiling with the BD Rhapsody™ system.
Single-Cell Labeling with BD® Flex Single-Cell Multiplexing Kits and BD® AbSeq Ab-Oligos
This protocol describes the use of BD® AbSeq Ab-Oligos (antibody-oligonucleotides) with the BD® Flex Single-Cell Multiplexing Kits (Cat. no. 633849-633852).
BD® AbSeq Immune Discovery Panel (IDP)
The IDP consists of 30 different specificities against major human immune markers combined in a single tube. Designed and optimized to work on the BD Rhapsody™ Single-Cell Analysis System, the IDP is tested to work seamlessly with BD Rhapsody Single-Cell RNA Assays and Multiplexing Kits. In this brochure, we also describe steps for reconstituting the lyophilized IDP and staining cells.
cDNA Synthesis Protocols
BD Rhapsody™ Express Single-Cell Analysis System
Single Cell Capture and cDNA Synthesis with the BD Rhapsody™ Express Single-Cell Analysis System
This protocol describes cell loading in the BD Rhapsody™ Cartridge and single cell capture with the BD Rhapsody™ Express Single-Cell Analysis system.
Single Cell Capture and cDNA Synthesis with the BD Rhapsody™ Single-Cell Analysis System
This protocol describes cell loading in the BD Rhapsody™ Cartridge and single cell capture with the BD Rhapsody™ Single-Cell Analysis system with the BD Rhapsody™ Scanner.
BD Rhapsody™ HT Single-Cell Analysis System
BD Rhapsody™ HT Xpress System Single-Cell Capture and cDNA Synthesis Protocol
This protocol describes cell loading in the BD Rhapsody™ 8-Lane Cartridge and single-cell capture with the BD Rhapsody™ HT Xpress System.
BD Rhapsody™ HT Single-Cell Analysis System Single-Cell Capture and cDNA Synthesis Protocol
This protocol describes cell loading in the BD Rhapsody™ 8-Lane Cartridge and single-cell capture with the BD Rhapsody™ HT Single-Cell Analysis System.
Sequencing Library Preparation Protocols
BD® AbSeq (Protein-only) Library Preparation Protocols (for AbSeq-based cell calling)
BD® AbSeq Library Preparation Protocol (for AbSeq-based cell calling)
This protocol enables users to use cell surface proteins to profile cells independently of mRNA. Included here are instructions for how to generate BD® AbSeq single-cell libraries using the BD Rhapsody™ Single-Cell Analysis system, or the BD Rhapsody™ Express Single-Cell Analysis system. Sequencing of libraries are completed on Illumina sequencers followed by data analysis that utilizes BD® AbSeq-based cell calling pipeline which identifies cells without mRNA information.
BD® AbSeq and Sample Tag Library Preparation Protocol (for AbSeq-based cell calling)
This protocol enables users to use cell surface proteins to profile cells independently of mRNA. Included here are instructions for how to generate BD® AbSeq single-cell libraries and Sample Tag libraries using the BD Rhapsody™ Single-Cell Analysis system, or the BD Rhapsody™ Express Single-Cell Analysis system. Sequencing of libraries are completed on Illumina sequencers followed by data analysis that utilizes a BD protein cell calling pipeline which identifies cells without mRNA information.
Whole Transcriptome Analysis (WTA) Library Preparation Protocols
mRNA Whole Transcriptome Analysis (WTA) Library Preparation Protocol
This protocol provides instructions on creating a single cell whole transcriptome mRNA library after cell capture on the BD Rhapsody™ Single-Cell Analysis System or the BD Rhapsody™ Express Single-Cell Analysis System for sequencing on Illumina sequencers.
mRNA Whole Transcriptome Analysis (WTA) and AbSeq Library Preparation Protocol
This protocol provides instructions on creating single cell whole transcriptome mRNA and AbSeq protein detection libraries after cell capture on the BD Rhapsody™ Single-Cell Analysis System or the BD Rhapsody™ Express Single-Cell Analysis System for sequencing on Illumina sequencers.
mRNA Whole Transcriptome Analysis (WTA) and Sample Tag Library Preparation Protocol
This protocol provides instructions on creating single cell whole transcriptome mRNA and Sample Tag libraries after cell capture on the BD Rhapsody™ Single-Cell Analysis System or the BD Rhapsody™ Express Single-Cell Analysis System for sequencing on Illumina sequencers.
mRNA Whole Transcriptome Analysis (WTA), AbSeq, and Sample Tag Library Preparation Protocol
This protocol provides instructions on creating single cell whole transcriptome mRNA, AbSeq, and Sample Tag libraries after cell capture on the BD Rhapsody™ Single-Cell Analysis System or the BD Rhapsody™ Express Single-Cell Analysis System for sequencing on Illumina sequencers.
Targeted Gene Expression Analysis Library Preparation Protocols
mRNA Targeted Library Preparation Protocol
This protocol provides instructions on creating a single cell mRNA library after cell capture on the BD Rhapsody™ Single-Cell Analysis system or the BD Rhapsody™ Express Single-Cell Analysis system for sequencing on Illumina sequencers.
mRNA Targeted and Sample Tag Library Preparation Protocol
This protocol provides instructions on creating single cell mRNA and Sample Tag libraries after cell capture on the BD Rhapsody™ Single-Cell Analysis system or the BD Rhapsody™ Express Single-Cell Analysis system for sequencing on Illumina sequencers.
mRNA Targeted and BD® AbSeq Library Preparation Protocol
This protocol provides instructions on creating mRNA and BD® AbSeq single cell libraries with the BD Rhapsody™ Single-Cell Analysis system or the BD Rhapsody™ Express Single-Cell Analysis system for sequencing on Illumina sequencers.
mRNA Targeted, BD® AbSeq, and Sample Tag Library Preparation Protocol
This protocol provides instructions on creating single cell mRNA, Sample Tag, and BD® AbSeq libraries with the BD Rhapsody™ Single-Cell Analysis system or the BD Rhapsody™ Express Single-Cell Analysis system for sequencing on Illumina sequencers.
BD Rhapsody™ ATAC-Seq Assay Library Preparation Protocols
ATAC-Seq Library Preparation Protocol
This protocol provides instructions on generating a single-cell ATAC library using the BD Rhapsody™ Single-Cell Analysis System.
ATAC-Seq + WTA Library Preparation Protocol
This protocol provides instructions on generating a single-cell ATAC library and whole transcriptome analysis (WTA) mRNA library with cell-captured beads using the BD Rhapsody™ Single-Cell Analysis System.
BD Rhapsody™ TCR/BCR Next Assay Library Preparation Protocols
Human TCR/BCR Next + Targeted mRNA Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, and BCR libraries.
Human TCR/BCR Next + Targeted mRNA + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Human TCR/BCR Next + Targeted mRNA + AbSeq Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and BD® AbSeq libraries.
Human TCR/BCR Next + Targeted mRNA + AbSeq + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, BD® AbSeq, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Human TCR/BCR Next + WTA Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, and BCR libraries.
Human TCR/BCR Next + WTA + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Human TCR/BCR Next + WTA + AbSeq Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and BD® AbSeq libraries.
Human TCR/BCR Next + WTA + AbSeq + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, BD® AbSeq, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before
capture on the BD Rhapsody™ system.
Mouse TCR/BCR Next + Targeted mRNA Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, and BCR libraries.
Mouse TCR/BCR Next + Targeted mRNA + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Mouse TCR/BCR Next + Targeted mRNA + AbSeq Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and BD® AbSeq libraries.
Mouse TCR/BCR Next + Targeted mRNA + AbSeq + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, BD® AbSeq, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Mouse TCR/BCR Next + WTA Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, and BCR libraries.
Mouse TCR/BCR Next + WTA + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Mouse TCR/BCR Next + WTA + AbSeq Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, and BD® AbSeq libraries.
Mouse TCR/BCR Next + WTA + AbSeq + Sample Tag Library Preparation Protocol
This protocol enables high throughput single-cell transcriptome and protein analysis alongside TCR and BCR profiling of individual cells captured on the BD Rhapsody™ system, providing instructions for amplifying Illumina-compatible single-cell barcoded mRNA, TCR, BCR, BD® AbSeq, and Sample Tag libraries. The addition of a Sample Tag library enables demultiplexing of samples that were pooled before capture on the BD Rhapsody™ system.
Bioinformatics Guides
BD Rhapsody™ Sequence Analysis Pipeline User’s Guide
This guide provides detailed instructions on how to setup and run the BD Rhapsody™ Sequence Analysis Pipeline on the Seven Bridges Genomics platform or on a local server installation.
SeqGeq™ Documentation
SeqGeq was built by and for scientists, with powerful data reduction tools to help you discover, cluster, and subset populations and relationships. It also includes visualization tools—including heat mapping and easy-to-use report editors—to help you communicate your results. This site provides high-level descriptions of some of SeqGeq’s core functions.
Additional Resources
Processing Nuclei Samples on the BD Rhapsody™ Single-Cell Analysis System
In this technical bulletin, we show how isolated nuclei can be captured and amplified on the BD Rhapsody™ Single-Cell Analysis System.
Flow Cytometry Analysis of BD® AbSeq Ab-Oligo and Sample Tag Expression
The purpose of this protocol is to provide detailed instructions on how to use single-color flow cytometry to analyze expression of oligonucleotide-conjugated antibodies with a poly(A) tail. This protocol allows rapid analysis of the expression of protein targets in the sample of interest when labelling cells with Sample Tags from the BD® Single-Cell Multiplexing Kit or with BD® AbSeq Ab-Oligos. This protocol is used prior to cDNA Synthesis