SmartFlare^(TM) is a reliable method for assessing mRNA expression in single neural stem cells
作者机构：Department of Biomedical SciencesUniversity of CagliariMonserrato 09042CagliariItaly Laboratory of Stem Cells&Restorative NeurologyLund Stem Cell CenterLund UniversityLund SE-22184LundSweden Institute of NeuroscienceMedical University of InnsbruckInnsbruck 6020Austria
出 版 物：《World Journal of Stem Cells》 (世界干细胞杂志（英文版）（电子版）)
年 卷 期：2021年第13卷第12期
页 面：1918-1927页
核心收录：
学科分类：0710[理学-生物学] 07[理学] 071006[理学-神经生物学]
基 金：the"Fondo Integrativo per la Ricerca"(FIR)of the University of Cagliari Italy
主 题：mRNA detection SmartFlareTM NanoFlare Live staining Nanotechnology Neural stem cell genes
摘 要：BACKGROUND One of the most challenging tasks of modern biology concerns the real-time tracking and quantification of mRNA expression in living *** this matter,a novel platform called SmartFlare^(TM) has taken advantage of fluorophore-linked nanoconstructs for targeting RNA *** fluorescence emission does not account for the spatial mRNA distribution,NanoFlare technology has grown a range of theranostic applications starting from detecting biomarkers related to diseases,such as cancer,neurodegenerative pathologies or embryonic developmental *** To investigate the potential of SmartFlare^(TM) in determining time-dependent mRNA expression of prominin 1(CD133)and octamer-binding transcription factor 4(OCT4)in single living cells through *** Brain fragments from the striatum of aborted human fetuses aged 8 wk postconception were processed to obtain *** the in vitro differentiation,neurospheres were gently dissociated with Accutase *** cells were resuspended in a basic medium enriched with fetal bovine serum,plated on poly-L-lysine-coated glass coverslips,and grown in a lapse of time from 1 to 4 *** cell mRNA detection was performed using SmartFlare^(TM) probes(CD133,Oct4,Actin,and Scramble).All the samples were incubated at 37°C for 24 *** nuclear staining,Hoechst 33342 was ***^(TM) CD133-and OCT4-specific fluorescence signal was assessed using a semiquantitative visual approach,taking into account the fluorescence intensity and the number of labeled *** In agreement with previous PCR experiments,a unique expression trend was observed for CD133 and OCT4 genes until 7 d in vitro(DIV).Fluorescence resulted in a mixture of diffuse cytoplasmic and spotted-like pattern,also detectable in the contacting neural *** 15 to 30 DIV,only few cells showed a scattered fluorescent pattern,in line with the differentiation progression and coherent with mRNA downregulation of
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