Abstract
Abstract A variety of RNA analysis technologies is available for the detection of RNA transcripts from bulk cell populations. However, the techniques for RNA detection from single cells have been limited due to lack of sensitivity in detecting specific mRNAs present at low abundance in individual cells. By designing novel target-specific probes and adapting an in situ signal amplification method based on the RNAscope® detection platform, we demonstrate the specific and sensitive detection of intracellular RNAs in single cells by flow cytometry (RNAFlow). This method had sufficient sensitivity to distinguish cells containing low abundance RNA transcript. Furthermore, multiple distinct RNA targets were simultaneously detected with a high specificity in single cells without interference. The method can quantify the frequency of cells expressing specific RNA as well as the number of RNA copies in each expression-positive cell. We will present quantitative data to characterize RNA target-specific signal detection, demonstrated in applications such as HIV infection and immune activation. The RNAFlow assay, independently or as combined assay with co-protein detection, represents a valuable research tool for the specific and sensitive detection of multiple RNA transcripts in individual cells in heterogeneous biological specimens. Overall, this method will be useful for the analysis of functionally important RNA species from single cells, even at very low copy numbers.
Published Version
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