Expression profiling by high throughput sequencing Other
We developed a scalable assay permitting the simultaneous quantification of hundreds of proteins and the full transcriptome in thousands of individual cells from samples where cell number is limiting. The RNA Expression and Protein Sequencing assay (REAP-seq) uses DNA-labeled antibodies to permit DNA sequencing of both mRNA and antibodies in a single workflow using droplet microfluidics. We describe the development and validation of REAP-seq in human PBMCs, and use the assay to assess the costimulatory effects of an anti-CD27 agonist antibody in naïve CD8+ lymphocytes. Protein quantification using REAP-seq was more sensitive than parallel mRNA measurements and differentiated cell states with fewer analytes. Unbiased profiling of single cells without prior selection enabled identification and characterization of an unexpected cell type that would have been missed with population-averaged profiling methodologies.
The RNA Expression and Protein Sequencing assay (REAP-seq) which uses DNA-labeled antibodies to permit DNA sequencing of both mRNA and antibodies in a single workflow using droplet microfluidics was developed and validated in human PBMCs and naïve CD8+ T cells that were costimulated with either 1) aCD3 and aCD28 or 2) aCD3, aCD28 and aCD27.