Abstract

Multi-analyte Analysis in Single-cells using qPCR

Anders Stahlberg, Senior Scientist, University of Gothenberg

The single cell represents the basic unit of all organisms. Most investigations have been performed on large cell populations, but understanding cell dynamics and heterogeneity requires single-cell analysis. Current methods for single-cell analysis generally can detect only one class of analytes. Reverse transcription and the proximity ligation assay were coupled with quantitative PCR and used to quantify any combination of DNA, mRNAs, microRNAs (miRNAs), noncoding RNAs (ncRNAs), and proteins from the same single cell. The method has been demonstrated on transiently transfected human cells to determine the intracellular concentrations of plasmids, their transcribed mRNAs, translated proteins, and downstream RNA targets. This method is compatible with most cell-sampling approaches, and generates output for the same parameter for all measured analytes, a feature facilitating comparative data analysis. This approach should open up new avenues in molecular diagnostics for detailed correlation studies of multiple and different classes of analytes at the single-cell level. We will also discuss how single-cell gene expression data can be used to gain detailed information about cell types and cell states.