Enhanced mRNA FISH with compact quantum dots

Abstract

Fluorescence in situ hybridization (FISH) is the primary technology used to image and count mRNA in single cells, but applications of the technique are limited by photophysical shortcomings of organic dyes. Inorganic quantum dots (QDs) can overcome these problems but years of development have not yielded viable QD-FISH probes. Here we report that macromolecular size thresholds limit mRNA labeling in cells, and that a new generation of compact QDs produces accurate mRNA counts. Compared with dyes, compact QD probes provide exceptional photostability and more robust transcript quantification due to enhanced brightness. New spectrally engineered QDs also allow quantification of multiple distinct mRNA transcripts at the single-molecule level in individual cells. We expect that QD-FISH will particularly benefit high-resolution gene expression studies in three dimensional biological specimens for which quantification and multiplexing are major challenges.

Dye‐Loaded Nanoemulsions: Biomimetic Fluorescent Nanocarriers for Bioimaging and Nanomedicine

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Efficient base editing in G/C-rich regions to model androgen insensitivity syndrome

“…Most disease-associated genomic mutations are base substitutions and approximately half of pathogenic human single nucleotide polymorphisms (SNPs) are related to C-to-T substitutions in the ClinVar database. 1 Base editors (BEs), which combine Cas9-D10A nickase and APOBEC (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like) or AID (activation-induced deaminase) cytidine deaminase family members, 2 have been successfully applied to mediate C-to-T conversion in vitro and in vivo, 3 providing a powerful tool to model or repair diseaserelated human SNPs. Yet, the editing scope of BE3 was limited by the low editing efficiency at GpC dinucleotides and/or in regions with high CpG methylation levels.…”

Ultra-bright Raman dots for multiplexed optical imaging

“…The interior of cells appeared as porous media with a throat size of ~20 nm 21 . Imaging with quantum dots of various sizes also showed dramatic signal decrease as the hydrodynamic diameter increased 22 , 23 . Importantly, these two factors are inter-related.…”