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Versatile Multiplex Endogenous RNA Detection with Simultaneous Signal Normalization Using Mesoporous Silica Nanoquenchers

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posted on 15.12.2020, 17:03 by Peiyan Yuan, Xin Mao, Si Si Liew, Shuang Wu, Yi Huang, Lin Li, Shao Q. Yao
Detection of endogenous tumor-related RNA is vital for cancer diagnostics. Despite advancements made, live-cell RNA detection still faces numerous problems, such as low signal output and cell-to-cell variations arising from differences in probe uptake. To address these issues, we designed a versatile and highly sensitive mRNA/miRNA nanosensor featuring, for the first time, signal amplification and in-built signal normalization. Using dye-loaded mesoporous silica nanoquenchers (qMSNs) capped with target-corresponding antisense oligos (ASOs), direct fluorescence “Turn-ON” with signal amplification was achieved upon target binding. By readily varying the capping ASOs as well as cargo dyes, a suite of RNA nanosensors for multiplex target detection could be easily prepared. Further modification of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA-responsive molecular beacons (MBs) onto our nanosensor enabled dual detection of target RNA and GAPDH mRNA, allowing for target signal normalization using GAPDH as a reference. We demonstrated that this newly developed nanosensor could successfully differentiate between noncancer and cancer cells, as well as accurately monitor the relative expression levels of multiple tumor-related RNAs simultaneously in different cancer cell lines, with a high degree of specificity and sensitivity, functioning as a noninvasive “qPCR mimic” imaging tool in live cells.

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