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Methods Mol Biol. 2011;714:3-13. doi: 10.1007/978-1-61779-005-8_1.

Single molecule imaging of RNA in situ.

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Public Health Research Institute Center, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, Newark, NJ, USA.


This protocol describes a method to image individual mRNA molecules in situ. About 50 oligonucleotides complementary to different regions of a target mRNA species are used simultaneously. Each probe is labeled with a single fluorescent moiety. When these probes bind to their target, each mRNA molecule becomes so intensely fluorescent that it can be seen as a fine fluorescent spot. Several different mRNA species can be detected in multiplex imaging using differently colored probe sets for each species. An automated image-processing program is used to count the number of mRNA molecules of each species that are expressed in each cell, thus yielding single-cell gene expression profiles.

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