Scientific Techniques Using Antibody-Oligonucleotide Conjugates

Antibody-Oligonucleotide Conjugate Technology Development

Antibody-oligonucleotide conjugation has revolutionized molecular detection since the 1992 introduction of Immuno-PCR, achieving femtomolar sensitivity through DNA amplification.
Proximity Ligation Assay (PLA) marked a crucial technological breakthrough in 2004, enabling detection of proteins in a “sandwich” format with 2 antibodies to improve specificity + being an early “no wash” assay.
Technical evolution from streptavidin-biotin bridges to direct conjugation chemistry has dramatically improved signal-to-noise ratios while reducing sample requirements.
Integration of next-generation sequencing as a readout methodology has supercharged these techniques, enabling massive multiplexing capabilities impossible with traditional PCR detection.
This technological foundation continues to drive scientific progress, demonstrating how innovative tools enable new scientific possibilities. “Progress depends on the interplay of techniques, discoveries, and ideas, probably in that order." - Sydney Brenner.

Intro and purpose

This summary table catalogs key scientific techniques employing antibody-oligonucleotide conjugates, spanning from early detection methods to advanced spatial and multi-omics technologies.
We aim to continuously update this resource as new methods emerge and existing techniques evolve.
Future versions will include expanded technical details of oligo design, comparative sensitivity/specificity metrics, and emerging commercial applications.

Comprehensive Classification Table

Technique	Primary Purpose	Mechanism	Key Applications	Readout Method	Sample Type	First Published	Notable References
IN-DEPTH	Multiplexed spatial analysis of proteins and RNAs	Antibody-oligo conjugates for proteomic mapping + RNA probes for transcriptomic analysis	High-resolution spatial mapping of multiple proteins and the transcriptome	Fluorescence imaging + NGS sequencing	Tissue sections	2024	Yeo et al, bioRxiv 2024
MERFISH-Protein	Spatially resolved, multiplexed protein detection	Adaptation of MERFISH for protein detection using antibody-oligo conjugates	High-resolution spatial mapping of multiple proteins	Fluorescence imaging	Tissue sections	2023	Wang et al, SFN 2023
Digital Spatial Profiling (DSP)	Multiplexed spatial quantification of proteins and RNAs	UV-cleavable oligo tags on antibodies or RNA probes; oligos released and quantified	Spatial profiling of up to 44 proteins and 1,412 genes in FFPE tissues	NGS sequencing	FFPE tissue sections	2019	Merritt et al, Nature Biotechnology (2019)
Immuno-SABER	Signal amplification for spatial protein detection	DNA concatemers for signal amplification; DNA-barcoded antibodies imaged via iterative fluorescence cycles	High-resolution spatial mapping of proteins in tissues	Fluorescence imaging	Tissue sections	2019	Saka et al, Nature Biotechnology (2019)
Immuno-PCR (IPCR)	Increased sensitivity vs ELISA	Biotinylated DNA + streptavidin-Protein A + Antibody-PCR amplification	Protein detection at femtomolar/attomolar levels; pathogen detection	PCR-Based	Solution-Phase	1992	Sano et al., Science (1992)
Proximity Ligation Assay (PLA)	Protein detection	Antibody pairs with attached oligos brought into proximity allow DNA ligation and amplification	Protein-protein interactions; post-translational modifications; multiplexed detection using qPCR in 2007	PCR-Based	Solution-Phase	2006	Gullberg et al (2004); Fredriksson et al (2007)
In situ PLA	Visualization of proteins and protein-protein interactions in tissue	Version of PLA. Rolling Circle Amplification then fluorescent oligos for visualization	Tissue-specific protein interactions; cancer biomarkers	Microscopy-Based	Fixed Cells/Tissues	2006	Söderberg et al., Nature Methods (2006); Klaesson et al., Scientific Reports (2018)
Proximity Extension Assay (PEA)	Multiplex protein detection	Paired antibodies with DNA tags but using polymerase not ligase	Biomarker discovery; proteomics; multiplexed assays	PCR-Based; NGS-Based	Solution-Phase	2011	Lundberg et al 2011; Assarsson et al., PLoS One (2014)
DNA-PAINT	Super-resolution imaging	Transient binding of fluorescent oligos to antibody-coupled DNA docking strands	Nanoscale imaging; subcellular structures; protein complexes	Microscopy-Based	Fixed Cells/Tissues	2010	Schnitzbauer et al., Nature Protocols (2017)
CODEX (CO-Detection by indEXing)	Highly multiplexed imaging	Sequential imaging using DNA-barcoded antibodies and cyclic nucleotide exchange	Spatial proteomics; tumor microenvironment; cell atlases	Microscopy-Based	Spatially-Resolved Tissue	2018	Goltsev et al., Cell (2018)
CITE-seq/REAP-seq	Simultaneous protein-RNA profiling	Antibody-oligos detected alongside transcriptome in single cells	Cell atlas building; immunology; cancer heterogeneity	NGS-Based	Single-Cell Suspensions	2017	Stoeckius et al., Nature Methods (2017); Peterson et al., Nature Biotechnology (2017)
Spatial-CITE-seq	Spatial protein-RNA co-mapping	Integration of spatial transcriptomics with CITE-seq	Spatial cell typing; tissue microenvironment; developmental biology	NGS-Based	Resolved Tissue	2023	Liu et al., Nature Biotechnology (2023)

Achieving batch-to-batch consistency with Antibody Oligonucleotide Conjugates (AOCs)