BD® SMK Staining Protocols

Single Cell Labeling

• Single Cell Labelling with 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. Every antibody-oligo in the BD® Human Single-Cell Multiplexing Kit, referred to as a Sample Tag, has a unique sample oligo barcode conjugated to a human universal antibody and every Sample Tag in the BD® Mouse Immune Single-Cell Multiplexing Kit is conjugated to an Anti-Mouse CD45, Clone 30-F11 antibody. Up to 12 samples can be labelled and pooled prior to single cell capture with the BD Rhapsody™ Single-Cell Analysis system.
  • Doc ID: 23-21340(02)
  • Revision date: 11/2022
• Single Cell Labelling with BD® Single-Cell Multiplexing Kits and BD® AbSeq Ab-Oligos (1 to 40 plex)
  • This protocol describes the use of BD AbSeq Ab-Oligos (antibody-oligonucleotides)(1 plex to 40 plex) with BD® Single-Cell Multiplexing Kits.
  • Doc ID: 23-21339(02)
  • Revision date: 11/2022
• Single-Cell Labeling with BD® Single-Cell Multiplexing Kits and BD® AbSeq Ab-Oligos (41 to 100 plex)
  • This protocol describes the use of BD AbSeq Ab-Oligos (antibody-oligonucleotides)(41 plex to 100 plex) with BD® Single-Cell Multiplexing Kits.
  • Doc ID: 23-22354(02)
  • Revision date: 11/2022

If you are using Custom BD Single- Cell Multiplexing Set Rat Anti-Mouse MHC-H2 Class I (M1/42), follow this protocol

• For tagging your samples of interest with the customs Rat Anti Mouse MHC-H2 Class I (M1/42) SMK set, first, resuspend 20,000 – 1 million cells in 180 μl BD Pharmingen Stain Buffer (FBS) (Cat. No. 554656) in a clean tube for each of your samples to be tagged. We recommend staining in a 5 ml round bottom tubes.
• Next, add one test worth (20µl) of SMK reagent from a sample tag vial to each sample (up to 12 individual samples can be tagged using the 12 samples tags provided).
• Pipet-mix. Take care to note the Sample Tag number for each individual sample being stained in parallel.
• Then proceed to follow the incubation and washing conditions outlined in the Single Cell Labelling with the protocol called Single Cell Labelling with BD® Single-Cell Multiplexing Kits

These cell labelling protocols are followed by cell capture and then downstream library prep protocols for WTA, Targeted or VDJ(TCR/BCR) analysis.

If you are interested in an AbSeq only readout with Sample Multiplexing(without the mRNA readout), follow this library preparation protocol after single cell labelling

• BD® AbSeq and Sample Tag Library Preparation Protocol
  • 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. 
  • Doc ID : 23-24228(01)
  • Revision date : 12/2022

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