Multiplexed, targeted profiling of single-cell proteomes and transcriptomes in a single reaction

Area of Science:

• Biochemistry
• Molecular Biology
• Genomics

Background:

• Single-cell analysis is crucial for understanding cellular heterogeneity.
• Simultaneous measurement of protein and RNA provides a more comprehensive view of cellular states.
• Existing methods for coupled protein and RNA detection can be complex and lack scalability.

Purpose of the Study:

• To present a scalable, integrated strategy for coupled protein and RNA detection from single cells.
• To profile the transcriptomic and proteomic response of a human breast adenocarcinoma cell line to chemical perturbation.
• To demonstrate the synergies enabled by simultaneous measurement of single-cell protein and RNA abundances.

Main Methods:

• Leveraging DNA polymerase activity of reverse transcriptase for simultaneous proximity extension assays and cDNA synthesis.
• Utilizing the Fluidigm C1 system for single-cell processing.
• Benchmarking against in situ hybridizations, immunofluorescence staining, recombinant proteins, ERCC Spike-Ins, and population lysate dilutions.

Main Results:

• Demonstrated a scalable and integrated approach for coupled protein and RNA detection.
• Successfully profiled the response of a human breast adenocarcinoma cell line to chemical perturbation.
• Showcased synergies from simultaneous measurement of single-cell protein and RNA, validated against established methods.

Conclusions:

• The developed generalizable approach enables highly-multiplexed single-cell analyses.
• Simultaneous protein and RNA detection provides valuable molecular metadata for deeper biological insights.
• This strategy holds significant potential for advancing single-cell research across various disciplines.