Simultaneous single-cell profiling of the transcriptome and proteome

Transcriptomic and proteomic measurements from the same single cell provide complementary information that cannot be inferred from either modality alone, yet methods for the parallel recovery of both analyte classes from a single-cell lysate remain limited. Here, we describe a workflow in which individual cells are isolated by automated dispensing into a minimal, MS-compatible lysis volume, followed by sequential mRNA capture and protein supernatant recovery, prior to independent downstream processing. The method is compatible with standard library preparation and data-independent acquisition proteomics pipelines and requires no dedicated instrumentation beyond a single-cell dispensing platform.

We evaluated workflow performance on 67 single cells across 3 iBlastoids. Transcriptomic sequencing detected a median of 5375 genes per cell, and proteomic analysis identified a median of 2123 protein groups per cell across two mass spectrometry platforms. Compared with a standalone single-cell proteomics protocol, incorporating the mRNA extraction step reduced median proteomic depth by approximately 11% (median 1,965 vs. 2,204 protein groups per cell), while mean percell identification remained comparable across workflows (1,790 vs. 1,775 protein groups per cell). Direct comparison of paired transcript and protein abundance yielded a median Spearman correlation of ρ ≈ 0.38; after correction for detection depth, the partial correlation was 0.067.