Table 1.

Selected examples of single-cell technologies that enable characterization of multiple features

	Technology	Throughput	Highlight	Reference
FC/microscopy & protein	scWB	Up to thousands of single cells	Combination of microwells and PAGE gel for protein detection based on mass or/and pI Can detect up to 11 proteins on the same cells by antibody stripping/re-probing Compatible with FACS sorting or cell imaging as pre-characterization Re-probing archival sample is possible	[41–46]
Protein&mRNA	FC	Millions	Fusion of Flow-FISH & ICS Quantification of mRNA + protein of three cytokines simultaneously (IFN-γ, IL-2, and TNF-α)	[48]
	MC	Millions	PLAYR Relies on MC (mass tag) or FC (fluorophore) for protein (antibody) and transcript (oligonucleotides) read-out The multiplexing capacity is determined by the available tags (~40 MC)	[51]
	Single-cell PCR	96 (using Fluidigm™ C1)	Leverages the DNA polymerase activity of reverse transcriptase to simultaneously perform proximity extension assays and complementary DNA synthesis in the same reaction Compatible with scRNA-seq platform Demonstrated detection of 96 RNA + 38 proteins	[58]
scRNA-seq&TCR-seq	TraCeR, scTCRseq, VDJPuzzle, TRAPeS	Depends on throughput of scRNA- seq	Extract TCR information from scRNA-seq results Provide both transcriptional profiling and clonality of single T-cell	[65–68]
scRNA-seq	scRNA-seq	Thousands of single cells	CITE-seq (10 surface proteins), REAP-seq (82 proteins) Oligonucleotide with poly A tail as unique antibody barcode Use reverse transcriptase as DNA polymerase to extend antibody barcode and reverse transcription of mRNA simultaneously Compatible with current scRNA-seq platform	[69,70]
	Abseq	>10,000 cells	Detection protein via DNA-labeled antibody to increase multiplexing capacity Each antibody also has UMI sequence for PCR- bias correction Compatible with current scRNA-seq platform Theoretical limit of detected protein is determined by sequencing depth and availability of antibody	[71]
	Seq-Well	~15,000 cells	Co-capture cells and transcripts-capture beads within individual microwells On-chip lysis, reverse transcription in bulk Compatible with on-chip imaging cytometry Transcriptomic profiling is done by scRNA-seq	[72]
Integration: cell-cell interaction, protein, etc	TIMING	20,000 cells	Co-culture lymphocyte and target cell on the same individual microwell Time-lapse microscopy live cell imaging Integrated with real-time cytokine secretion and cell retrieval for gene expression profiling	[77–81]
	Droplet	~1,000 events	Similar to TIMING Co-encapsulate two types of cells within a droplet Droplet docking into individual microwell	[82]
	SCBC	Up to several thousands of cells	Single cells or cell pairs are isolated in individual microchambers Cell-cell interaction can be investigated Detection antibody coated surface is detachable for analysis Up to 45-plex protein detection including secreted proteins	[84–86]
	BOBarray	Several thousands of cells	Similar to SCBC, but use antibody-coated beads as protein sensor Miniaturized device achieved by combination of bead size and fluorophore combination: 4 bead size × 3 color =12-plex	[87]