In situ visualization of messenger RNA for basic fibroblast growth factor in living cells.
We examined whether messenger RNA for basic fibroblast growth factor (bFGF) could be visualized specifically by a fluorescent probe in living cells. A 15-nucleotide-long antisense or sense sequence for human bFGF was sandwiched between two complementary 5-nucleotide-long arm sequences. A fluorophore, 5-(2'-aminoethyl)aminonaphthalene-1-sulfonic acid (EDANS), was joined to the 5'-terminal phosphate, while 4-(4'-dimethylaminophenylazo)benzoic acid, quencher for EDANS, was joined to the 3'-terminal hydroxyl group. The probe emitted blue fluorescence only upon hybridization with the complementary 18-nucleotide-long sequence under ultraviolet light. The antisense or sense probe carried with liposome was delivered to human cells, trabecular cells of the eye, in a glass-bottom culture dish placed on the stage of an inverted microscope. Cells with the antisense probe did, but not with the sense probe, show blue fluorescence under ultraviolet light. The present study opens a way to measure the changing levels of a specific messenger RNA in living cells.
Matsuo
T
T
Department of Ophthalmology, Okayama University Medical School, Japan.
D001483:Base Sequence; D002478:Cells, Cultured; D016222:Fibroblast Growth Factor 2 / Q000235:genetics*; D005456:Fluorescent Dyes / Q000138:chemical synthesis; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D015336:Molecular Probe Techniques; D015345:Oligonucleotide Probes / Q000138:chemical synthesis / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism*; D014129:Trabecular Meshwork / Q000166:cytology / Q000378:metabolism
D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D016222:Fibroblast Growth Factor 2