Bromodeoxyuridine-labeled oligonucleotides as tools for oligonucleotide uptake studies.
The mechanisms by which various oligonucleotides (ODNs) and their analogs enter cells are not fully understood. A common technique used in studies on cellular uptake of ODNs is their conjugation with fluorochromes. However, fluorescently labeled ODNs may vary from the parent compounds in charge and hydrophilicity, and they may interact differently with some components of cellular membranes. In this report, we present an alternative method based on the immunofluorescent detection of ODNs with incorporated 5-bromo-2'-deoxyuridine (BrdUrd). Localization of BrdUrd-modified ODNs has been achieved using FITC-labeled anti-BrdUrd antibodies. This technique allowed determination of the differences in cellular uptake of phosphodiester (PO) and phosphorothioate (PS) ODNs and their derivatives conjugated with cholesterol and menthol. The immunocytochemical method also has shown that the cellular uptake of some ODNs may be influenced by specific sequences that are responsible for the formation of higher-order structures.
Maszewska
Maria
M
Department of Bioorganic Chemistry, Centre of Molecular and Macromolecular Studies, Polish Academy of Sciences, 90-363 Lódź, Sienkiewicza 112, Poland. maszewsk@bio.cbmm.lodz.pl
Kobylańska
Anna
A
Gendaszewska-Darmach
Edyta
E
Koziołkiewicz
Maria
M
D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D001973:Bromodeoxyuridine / Q000032:analysis* / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000378:metabolism*; D013194:Staining and Labeling
D009841:Oligonucleotides; D001973:Bromodeoxyuridine